C1q antibodies and uses thereof

Abstract

The present disclosure concerns isolated polypeptides comprising an antibody variable domain that specifically binds to human solid-phase complement component C1q. Such polypeptides may be used as a medicament, for example in the treatment of a disease or disorder that would benefit from either enhancement or reduction of the complement cascade. Corresponding conjugates, nucleic acids, cells, pharmaceutical compositions, kits, and uses are also described.

Description

[0001] C1Q ANTIBODIES AND USES THEREOF

[0002] Field of the invention

[0003] The present disclosure concerns isolated polypeptides comprising an antibody variable domain that specifically binds to human solid-phase complement component C1 q. Such polypeptides may be used as a medicament, for example in the treatment of a disease or disorder that would benefit from either enhancement or reduction of the complement cascade. Corresponding conjugates, nucleic acids, cells, pharmaceutical compositions, kits and uses are also described.

[0004] Background of the invention

[0005] Complement component 1q (C1 q) is a key protein of the complement system and its traditionally accepted role is to serve as a recognition molecule in the classical pathway of complement activation. C1q is mainly produced by myeloid-derived immune cells. It recognizes antigen-antibody complexes, as well as various other molecular targets. Binding of soluble (fluid-phase) C1q to its target induces a conformational change, and the resultant bound (solid-phase) C1 q triggers activation of complement via associated C1 r and C1 s proteases. Depending on the circumstances, C1q induced activation of complement may be advantageous (for example in response to infection or cancer), or it may have deleterious effects (for example in auto-immune disease or during transplant rejection).

[0006] Antibodies that bind to C1q or fluid phase C1 q are known, see for example Trouw et al.: “AntiCi q autoantibodies deposit in glomeruli but are only pathogenic in combination with glomerular C1 q- containing immune complexes”, Journal of Clinical Investigation, 2004. However, preferential binding of C1 q-binding monoclonal antibodies to solid-phase C1q is not known. Fluid-phase C1q-binding antibodies are used to systemically dampen down the immune system in situations where excessive C1 q induced complement activation is undesirable. Such antibodies may therefore be used as part of a treatment regimen for patients with auto-immune disease. However, these treatments also make the patient much more vulnerable to infections. In this setting the anti-C1q binds to fluid-phase C1q in the circulation and mediates its clearance. In addition, some anti-C1q autoantibodies have also been shown to trigger and enhance pathological effects. Alternative treatments that are specific for the autoimmune disease of interest are therefore generally preferred. However, although such treatments are more targeted, they do not have broad applicability. of the invention

[0007] The inventors have developed a new strategy for treating diseases or disorders that would benefit from either enhancement or reduction of the complement cascade. By providing a polypeptide (e.g. antibody) that specifically targets and binds to solid-phase C1q, the invention specifically targets C1 q that is bound to its target ligand, and thus can be used to either induce or inhibit the complement cascade. This provides a completely new treatment modality for patients with diseases or disorders that would benefit from either enhancement or reduction of the complement cascade exactly at the location where C1 q is binding to its targets. The invention has particular utility in treating auto-immunity. The inventors have identified that targeting solid-phase C1 q (as opposed to fluid-phase C1 q) provides a means for specifically targeting C1q that is contributing to tissue damage in auto-immunity without depleting circulating C1q levels. This avoids the problems associated with targeting fluid-phase C1q (i.e., it avoids inducing susceptibility to infections), and maintains broad applicability (such that the therapy can be used generically to treat several different auto-immune diseases).

[0008] In order to implement the invention, the inventors have identified several C1q antibodies that are specific for solid-phase C1 q only (see examples below). Although the invention may be implemented using one or more of these antibodies, the invention is not limited to use of these specific antibodies and extends to any polypeptide or antibody that specifically binds to solid-phase C1q.

[0009] Advantageously, a polypeptide according to the invention will not substantially be affected by fluid-phase C1q present in the systemic circulation of a subject. Circulating levels of C1 q in human blood or serum can be of from 100 to 500 pg / ml and present a major potential sink for any non-specific solid-phase C1q binding polypeptides or antibodies. Thus, specific targeting of complement pathway enhancement or inhibition is possible using the polypeptides according to the invention while sparing the systemic complement pool.

[0010] Furthermore, by specifically targeting solid-phase C1 q bound to one of its natural ligands (such as but not limited to complexed IgG, surface-bound IgM, C-reactive protein, HIV-1 , phosphatidylserine, HTLV-1 , myelin debris, amyloid fibrils, post-translationally modified proteins, and apoptotic cells) a lower dose of medication may be used (compared to a medication that targets fluid-phase and solidphase C1q).

[0011] The object of the present invention is therefore to provide an isolated polypeptide comprising an antibody variable domain that specifically binds to human solid-phase complement component C1 q.

[0012] It is a further object to provide a polypeptide according to the invention, wherein the antibody variable domain comprises: a. a heavy chain variable domain (VH) comprising

[0013] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 1 ;

[0014] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 2;

[0015] (iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 3; and a light chain variable domain (VL) comprising

[0016] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 4;

[0017] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 5; (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 6; or b. a heavy chain variable domain (VH) comprising

[0018] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 7;

[0019] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 8;

[0020] (iii) a CDR-H3 comprising an amino acid sequence having at least 92% sequence identity to SEQ ID NO: 9; and a light chain variable domain (VL) comprising

[0021] (i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 10;

[0022] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 11 ;

[0023] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 12; or c. a heavy chain variable domain (VH) comprising

[0024] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 13;

[0025] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 14;

[0026] (iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 15; and a light chain variable domain (VL) comprising

[0027] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 16;

[0028] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 5;

[0029] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 17; or d. a heavy chain variable domain (VH) comprising

[0030] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 18;

[0031] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 19;

[0032] (iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 20; and a light chain variable domain (VL) comprising (i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;

[0033] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 22;

[0034] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 23; or e. a heavy chain variable domain (VH) comprising

[0035] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 24;

[0036] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 25;

[0037] (iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 26; and a light chain variable domain (VL) comprising

[0038] (i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;

[0039] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 22;

[0040] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 27; or f. a heavy chain variable domain (VH) comprising

[0041] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 28;

[0042] (ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 29;

[0043] (iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 30; and a light chain variable domain (VL) comprising

[0044] (i) a CDR-L1 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 31 ;

[0045] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 32;

[0046] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 33; or g. a heavy chain variable domain (VH) comprising

[0047] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 34; (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 35;

[0048] (iii) a CDR-H3 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 36; and a light chain variable domain (VL) comprising

[0049] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37;

[0050] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 38;

[0051] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 39; or h. a heavy chain variable domain (VH) comprising

[0052] (i) a CDR-H1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 40;

[0053] (ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 41 ;

[0054] (iii) a CDR-H3 comprising an amino acid sequence having at least 93% sequence identity to SEQ ID NO: 42; and a light chain variable domain (VL) comprising

[0055] (i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 43;

[0056] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 44;

[0057] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45; or i. a heavy chain variable domain (VH) comprising

[0058] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 46;

[0059] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 47;

[0060] (iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 48; and a light chain variable domain (VL) comprising

[0061] (i) a CDR-L1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 49;

[0062] (ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 50; (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 51.

[0063] It is yet a further object to provide a conjugate comprising a polypeptide according to the invention conjugated to or recombinantly fused to a diagnostic agent, detectable agent, or therapeutic agent.

[0064] It is yet a further object to provide an isolated nucleic acid sequence encoding a polypeptide or a conjugate according to the invention.

[0065] It is yet a further object to provide an isolated cell comprising a nucleic acid sequence according to the invention.

[0066] It is yet a further object to provide a pharmaceutical composition comprising a polypeptide or conjugate according to the invention, and a pharmaceutically acceptable excipient, adjuvant, diluent, or carrier.

[0067] It is yet a further object to provide a kit comprising a polypeptide or conjugate according to the invention, and instructions for using the polypeptide or conjugate to (i) treat or prevent infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in a subject in need of such treatment, or (ii) diagnose or predict the development of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in vivo in a subject.

[0068] It is yet a further object to provide a polypeptide or conjugate according to the invention, or a pharmaceutical composition according to the invention, for use as a medicament or a diagnostic.

[0069] It is yet a further object to provide a use of a polypeptide or conjugate according to the invention for detecting solid-phase C1 q.

[0070] It is yet a further object to provide a method of detecting solid-phase C1 q in a biological sample, comprising the steps of: a. contacting a polypeptide or conjugate according to the invention with the biological sample; and b. detecting the presence or absence of the polypeptide bound to solid-phase C1 q in the biological sample.

[0071] It is yet a further object to provide a method of detecting solid-phase C1q in vivo in a subject, comprising the steps of: a. administering a polypeptide or conjugate according to the invention, or a pharmaceutical composition according to the invention, to the subject; and b. detecting the presence or absence of the polypeptide bound to solid-phase C1 q in the subject. It is yet a further object to provide a method of treating a subject, comprising administering a therapeutically effective amount of a pharmaceutical composition according to the invention to a subject in need thereof.

[0072] Description of the Figures

[0073] Embodiments of the invention are further described hereinafter with reference to the accompanying drawings, in which:

[0074] Figure 1 shows binding of anti-C1q monoclonal antibodies.

[0075] Figures 2 and 3 show binding of anti-C1q monoclonal antibodies to solid-phase C1 q in the presence of fluid-phase C1q as a competitor.

[0076] Figure 4 shows binding of anti-C1q monoclonal antibodies to solid-phase C1 q in the presence of fluid-phase C1q as a competitor.

[0077] Figure 5 shows competition with fluid-phase purified C1 and C1q depleted serum for selected anti-C1 q mAbs 1 F5, 4F5 and mouse 4A4B11 .

[0078] Figure 6 shows that binding of anti-C1 q antibodies is inhibited by solid-phase C1 q but not fluidphase C1q.

[0079] Figure 7A shows binding of anti-C1q monoclonal antibodies to solid-phase C1 q. Figure 7B shows binding of anti-C1q mAbs to IgG opsonized cells as detected by flow cytometry. Figure 7C shows binding of anti-C1 q mAbs to necrotic cells as detected by flow cytometry.

[0080] Figure 8 shows Western blots of C1q in various states of protein folding, with detection by antiCi q mAbs.

[0081] Figure 9A shows binding of anti-C1 q monoclonal antibodies to distinct non-competing epitopes on solid-phase C1 q. Figure 9B shows anti-C1q mAbs binding to full C1 q, C1q Collagen Like Region (CLR), and C1 q (recombinant) globular head (gh) domains.

[0082] Figure 10 shows uptake or binding of IgM- and C1q-labelled beads by THP-1 cells differentiated into macrophages.

[0083] Figure 11 shows electron tomography images of C1 q bound by anti-C1 q mAb.

[0084] Figure 12 shows binding of antibodies from SLE patient plasma, healthy anti-C1q negative serum, a mix of anti-C1q mAbs or polyclonal Rabbit anti-C1q (DAKO; A0136) to linear 21-mer C1q peptides.

[0085] Figure 13 shows that anti-C1 q mAbs increase Fc receptor engagement, but not complement activation, on immune complexes.

[0086] Figure 14 shows that the epitopes targeted by the anti-C1 q mAbs constitute the same epitopes targeted by anti-C1 q autoantibodies present in sera from systemic lupus erythematosus (SLE) patients.

[0087] Various aspects of the invention are described in further detail below.

[0088] Detailed Description of the Invention The present invention relates to an isolated polypeptide comprising an antibody variable domain that specifically binds to human solid-phase complement component C1 q.

[0089] Polypeptides are described herein that comprise an antibody variable domain that specifically binds to human solid-phase complement component C1 q. As would be clear to a person of skill in the art, such polypeptides may be, for example, an antibody binding fragment of an antibody. For example, they may be an scFv. Such polypeptides may also be part of a protein (e.g. protein complex), such as an antibody. For simplicity, the terms “polypeptide” and “protein” are used interchangeably herein. The polypeptides / proteins may therefore be an antibody or antigen-binding fragment thereof. In a specific example, the polypeptides / proteins may be a whole antibody, bispecific antibody, diabody, triabody, tetrabody, minibody, nanobody, sdAb, scFv, di-scFv, bis-scFv, tri-scFv, scFv-Fc, Fab, Fab', F(ab')2, or a Fv.

[0090] Polypeptides are described herein that comprise an antibody variable domain. As would be clear to a person skilled in the art, the polypeptides may therefore comprise one or more antibody variable domains e.g. two antibody variable domains. Antibody variable domains (e.g. VH or VL) are well known in the art and are described elsewhere herein.

[0091] “Antibody variable domain” herein is understood to mean the part of the heavy or light chain regions of an antibody that is used for binding to an antigen (and the part which provides specificity to the antigen). The terms “antibody variable domain”, “variable domain” and “variable region” are used interchangeably herein. As described elsewhere herein the complementary-determining regions (CDR) of the heavy and light chain variable domains contribute to antigen binding and specificity the most. An antigen-binding site of an antibody is made up of six CDRs, three from each of the light and heavy chains and are therefore termed CDR-L1 , CDR-L2, CDR-L3, CDR-H1 , CDR-H2 and CDR-H3. The CDRs can be arranged non-consecutively. of

[0092] The antibody variable domains provided herein specifically bind to human solid-phase complement component C1q (also referred to as solid-phase C1q herein). In the context of binding, the terms “specific” and “specifically” are used herein interchangeably to indicate that other biomolecules do not significantly bind to the antibody variable domain that is specifically binding to the biomolecule of interest (human solid-phase C1 q). In other words, the antibody variable domains provided herein specifically bind to solid-phase C1 q such that they bind to solid-phase C1 q with greater affinity, avidity, more readily, and / or with greater duration than other biomolecules or targets. Specific binding to a first antigen (e.g., solid-phase C1 q) may or may not include specific binding to a second antigen, and therefore may or may not represent exclusive binding to a target antigen.

[0093] In some examples, the level of binding to a biomolecule other than human solid-phase C1q results in a negligible (e.g., not determinable) binding affinity by means of ELISA or an affinity determination. By “negligible binding” a binding is meant, which is at least about 85%, particularly at least about 90%, more particularly at least about 95%, even more particularly at least about 98%, but especially at least about 99% and up to 100% less than the binding to human solid-phase C1q. The binding affinity of an antibody variable domain with solid-phase C1q may be determined using a standard binding assay, such as surface plasmon resonance technique (BIAcore®, GE- Healthcare Uppsala, Sweden). The term "surface plasmon resonance," as used herein, refers to an optical phenomenon that allows for the analysis of real-time biospecific interactions by detection of alterations in protein concentrations within a biosensor matrix, for example using the BIAcore system (Pharmacia Biosensor AB, Uppsala, Sweden and Piscataway, N.J.). For further descriptions, see Jonsson, U., et al. (1993) Ann. Biol. Clin. 51 : 19-26; Jonsson, U., et al. (1991) Biotechniques 11 :620- 627; Johnsson, B., et al. (1995) J. Mol. Recognit. 8: 125-131 ; and Johnnson, B., et al. (1991) Anal. Biochem. 198:268-277.

[0094] For example, “specifically binding” in the context of the binding of an antibody to a predetermined antigen / epitope means binding with an affinity corresponding to a KD (equilibrium dissociation constant) of about 10-7M or less, such as about 10-8M or less, such as about 10-9M or less, about 10"10M or less, or about 10"11M or even less when determined for instance by surface plasmon resonance (SPR) technology in a BIAcore 3000 instrument using the antibody as the analyte (wherein a low KD indicates a high affinity). This term also means that the antibody binds to the predetermined antigen / epitope with an affinity corresponding to a KD that is at least ten-fold lower, such as at least 100 fold lower, for instance at least 1000 fold lower, such as at least 10,000 fold lower, for instance at least 100,000 fold lower than its affinity for binding to a non-specific antigen (e.g., bovine serum albumin, casein) otherthan the predetermined antigen ora closely-related antigen. The amount with which the affinity is lower is dependent on the KD of the antibody, so that when the KD of the antibody is very low (that is, the antibody is very specific and binds very well), then the amount with which the affinity for the antigen is lower than the affinity for a non-specific antigen may be at least 10,000 fold. The term “KD” as used herein, means the dissociation rate constant of a particular antibody-antigen interaction.

[0095] An antibody variable domain described herein may be specific for (i.e. , specifically bind to) an epitope within solid-phase C1q. As used herein the term “epitope” refers to a site on a target molecule (e.g., an antigen, such as a protein, for example of solid-phase C1q) to which an antibody variable domain (e.g., a binding protein such as an antibody or antibody fragment) binds. Epitopes are groupings of molecules such as amino acids or sugar side chains and usually have specific structural characteristics, as well as specific charge characteristics. A single antigen may have more than one epitope. Epitopes can be formed both from contiguous or adjacent noncontiguous residues (e.g., amino acid residues) of the target molecule. Epitopes formed from contiguous residues (e.g., amino acid residues) typically are also called linear epitopes. An epitope typically includes at least 5 and up to about 12 residues, mostly between 6 and 10 residues (e.g., amino acid residues). Epitopes may also be conformational (i.e., non-linear).

[0096] In the context of the present disclosure, an antibody variable domain that specifically binds to human solid-phase complement component C1q as described herein does not specifically bind to fluid-phase C1 q. In other words, the antibody variable domain binds to solid-phase C1 q with a greater affinity, avidity, more readily, and / or with greater duration than its binding to fluid-phase C1 q. Methods for measuring binding specificity are well known in the art. For example, as described in the experimental section herein under "ELISA for fluid phase C1 q inhibition to anti-C1q”, an ELISA Competition Assay may be used, further exemplified by figures 2-4.

[0097] In an example provided herein, preferably the polypeptide according to the invention comprises an antibody variable domain that specifically binds to human solid-phase complement component C1q in that in an in vitro competitive binding assay the polypeptide exhibits less than 25%, preferably less than 20%, 15%, 10%, or 5%, inhibition in binding to solid-phase C1q in the presence of an excess of fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluidphase C1 q. Preferably, the excess of fluid-phase C1 q is 20 pg / ml purified fluid-phase C1 q. Preferably, the solid-phase C1q is provided at a concentration of 10 pg / ml. Preferably, the degree of inhibition is determined at or near the saturation concentration of the polypeptide when binding to solid-phase C1q in the absence of fluid-phase C1q. The saturation concentration of a polypeptide can be defined as the minimum concentration of the polypeptide required for maximum binding to a target ligand such as solid-phase C1q. The saturation concentration can be suitably determined by a person skilled in the art.

[0098] Preferably, the in vitro competitive binding assay comprises an ELISA Competition Assay and in the ELISA Competition Assay, the polypeptide exhibits less than 25% inhibition in binding of solidphase C1q in the presence of 20 pg / ml purified fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1 q in the absence of fluid-phase C1 q. Preferably, the solid-phase C1 q is provided by coating an ELISA plate surface with 10 pg / ml C1q, preferably wherein the coating is performed overnight at 4°C and / or followed by blocking with PBS / 1 %BSA. Preferably, the degree of inhibition is determined at or near the saturation concentration of the polypeptide when binding to solid-phase C1q in the absence of fluid-phase C1q. The saturation concentration of a polypeptide can be defined as the minimum concentration of the polypeptide required for maximum binding to a target ligand such as solid-phase C1 q. The saturation concentration can be suitably determined by a person skilled in the art.

[0099] In this context, in one example, the antibody variable domains described herein specifically bind solid-phase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 25% (such as less than 24%, 23%, 22%, 21 %, or 20%) inhibition in binding of solid-phase C1q in the presence of 20 pg / ml purified fluid-phase C1 q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q.

[0100] For example, the antibody variable domains described herein may specifically bind solidphase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 20% (such as less than 19%, 18%, 17%, 16%, or 15%) inhibition in binding of solid-phase C1q in the presence of 20 pg / ml purified fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q. In a further example, the antibody variable domains described herein may specifically bind solid-phase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 15% (such as less than 14%, 13%, 12%, 11%, or 10%) inhibition in binding of solid-phase C1 q in the presence of 20 pg / ml purified fluid-phase C1 q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q.

[0101] Particularly, the antibody variable domains described herein may specifically bind solid-phase C1 q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 10% inhibition in binding of solid-phase C1q in the presence of 20 pg / ml purified fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluidphase C1q.

[0102] The percentage of inhibition is determined at a non-saturating, but sufficient concentration of the polypeptide, which preferably is a concentration in the linear part of a titration curve of different concentrations of the polypeptide between at least one saturating concentration and at least one concentration insufficient to detect binding to solid-phase C1q in an ELISA competition assay. Exemplary titration curves can be found in figures 2 and 3. Exemplary selections of concentrations in the linear part of the titration curves can be found in figure 4. The above method is well-known and, when performed by a person skilled in the art, will yield similar, reproducible results.

[0103] In a further example, the antibody variable domains described herein may specifically bind solid-phase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 50% inhibition in binding to solid-phase C1q in the presence of 25% Normal Human Serum (NHS) comprising about 200 pg / ml fluid-phase C1 q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q.

[0104] For example, the antibody variable domains described herein may specifically bind solidphase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 40% inhibition in binding to solid-phase C1q in the presence of 25% Normal Human Serum (NHS) comprising about 200 pg / ml fluid-phase C1 q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q.

[0105] In this context, the antibody variable domains described herein may specifically bind solidphase C1q such that, in an ELISA Competition Assay, a polypeptide comprising the antibody variable domain exhibits less than 30% inhibition in binding to solid-phase C1q in the presence of 25% Normal Human Serum (NHS) comprising about 200 pg / ml fluid-phase C1 q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q.

[0106] For the avoidance of doubt, as a means for measuring binding specificity, an ELISA Competition Assay using NHS is less preferred compared to using purified fluid-phase C1q as described above.

[0107] In one example, the antibody variable domains described herein may specifically bind solidphase C1q such that, in an ELISA Competition Assay, binding of a polypeptide (comprising the antibody variable domain) to solid-phase C1q is inhibited two-fold more by competition of the binding with solid-phase C1q compared to competition with fluid-phase C1q

[0108] Preferably, the polypeptide according to the invention has a dissociation constant (KD) for solid-phase C1q of less than about 3.0 x 10'8M, preferably of less than about 2.5 x 10'8M, more preferably of less than about 2.4 x 10-8M or less than about 2.35 x 10'8M. See for example, the dissociation constants (KD) for solid-phase C1q provided below in Table 2 for the antibodies exemplified herein. The KD is preferably determined by surface plasmon resonance, preferably using a Biacore T200.

[0109] The antibody variable domains described herein specifically bind to human solid-phase complement component C1 q. As used herein, the terms “complement component C1 q” and “C1q” are used interchangeably. C1q is a 400 kDa protein formed from 18 peptide chains; six A-chains, six B- chains, and six C-chains. Each chain contains a collagen-like region located near the N terminus and a C-terminal globular region. The A-, B-, and C-chains are arranged in the order A-C-B on chromosome 1. See for example Structure and activation of the C1 complex of complement: unraveling the puzzle. Gaboriaud C, Thielens NM, Gregory LA, Rossi V, Fontecilla-Camps JC, Arlaud GJ. Trends Immunol. 2004 Jul;25(7):368-73. Doi: 10.1016 / j.it.2004.04.008.

[0110] C1 q exists in free and bound forms. These distinct forms are recognizable, as binding of C1q to its ligand results in a conformational change in C1q which enables the proteases C1 r and C1s to become active. C1 q associates with C1 r and C1 s in order to yield the C1 complex (C1 qr2s2), the first component of the serum complement system.

[0111] ‘Solid-phase C1 q’ herein is understood to mean C1 q bound to at least one of its natural ligands such as but not limited to complexed IgG, surface-bound IgM, C-reactive protein, HIV-1 , phosphatidylserine, HTLV-1 , myelin debris, amyloid fibrils, post-translationally modified proteins, or apoptotic cells. See for example, Circulating C1q levels in health and disease, more than just a biomarker. Van de Bovenkamp FS, Dijkstra DJ, van Kooten C, Gelderman KA, Trouw LA. Mol Immunol. 2021 Nov 2;140:206-216. Accordingly, “solid-phase” C1 q may also be described as the “bound” form of C1 q, or C1 q bound to one of its natural ligands. The bound nature of C1q is often determined by detecting C1 q bound to a target. However, the tools currently used to detect such bound-C1 q only work appropriately after washing away all the unbound C1 q.

[0112] ‘Fluid-phase C1 q’ herein is understood to mean C1q that is not bound to at least one of its natural ligands, for example when C1 q is freely circulating in blood or serum. Accordingly, “fluid-phase” C1 q may also be described as the “free” form of C1 q, or soluble C1q. Methods for identifying circulating, fluid-phase C1q are well known in the art, for example, ELISA or turbidimetry. Fluid-phase C1 q may be C1 q without C1 r and C1 s or may be in the form of the C1 complex that does contain C1 r and C1 s as long as the C1 complex is not bound to at least one of the natural ligands.

[0113] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0114] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 1 ;

[0115] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 2;

[0116] (iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 3; and a light chain variable domain (VL) comprising

[0117] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 4;

[0118] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 5;

[0119] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 6.

[0120] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises:

[0121] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 1 , ora functional variant or functional fragment thereof;

[0122] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 2, ora functional variant or functional fragment thereof;

[0123] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 3, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0124] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 4, or a functional variant or functional fragment thereof;

[0125] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 5, ora functional variant or functional fragment thereof;

[0126] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 6, or a functional variant or functional fragment thereof.

[0127] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises:

[0128] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 1 ;

[0129] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 2;

[0130] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 3; and a light chain variable domain (VL) comprising

[0131] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 4;

[0132] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 5;

[0133] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 6. An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “2D2”.

[0134] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 63; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 59.

[0135] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 63; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 59.

[0136] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “2D2”.

[0137] In an alternative example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0138] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 7;

[0139] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 8;

[0140] (iii) a CDR-H3 comprising an amino acid sequence having at least 92% sequence identity to SEQ ID NO: 9; and a light chain variable domain (VL) comprising

[0141] (i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 10;

[0142] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 11 ;

[0143] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 12.

[0144] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0145] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 7, ora functional variant or functional fragment thereof;

[0146] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 8, ora functional variant or functional fragment thereof; (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 9, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0147] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 10, or a functional variant or functional fragment thereof;

[0148] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 11 , or a functional variant or functional fragment thereof;

[0149] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 12, or a functional variant or functional fragment thereof.

[0150] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0151] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 7;

[0152] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 8;

[0153] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 9; and a light chain variable domain (VL) comprising

[0154] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 10;

[0155] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 11 ;

[0156] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 12.

[0157] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “1 F4”.

[0158] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 68; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 56.

[0159] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 68; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 56.

[0160] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “1 F4”.

[0161] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0162] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 13; (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 14;

[0163] (iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 15; and a light chain variable domain (VL) comprising

[0164] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 16;

[0165] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 5;

[0166] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 17.

[0167] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0168] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 13, or a functional variant or functional fragment thereof;

[0169] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 14, or a functional variant or functional fragment thereof;

[0170] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 15, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0171] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 16, or a functional variant or functional fragment thereof;

[0172] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 5, ora functional variant or functional fragment thereof;

[0173] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 17, or a functional variant or functional fragment thereof.

[0174] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0175] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 13;

[0176] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 14;

[0177] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 15; and a light chain variable domain (VL) comprising

[0178] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 16;

[0179] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 5;

[0180] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 17. An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “4C11”.

[0181] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 65; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 61.

[0182] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 65; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 61 .

[0183] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “4C11”.

[0184] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0185] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 18;

[0186] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 19;

[0187] (iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 20; and a light chain variable domain (VL) comprising

[0188] (i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;

[0189] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 22;

[0190] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 23.

[0191] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0192] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 18, or a functional variant or functional fragment thereof;

[0193] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 19, or a functional variant or functional fragment thereof; (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 20, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0194] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 21 , or a functional variant or functional fragment thereof;

[0195] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 22, or a functional variant or functional fragment thereof;

[0196] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 23, or a functional variant or functional fragment thereof.

[0197] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0198] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 18;

[0199] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 19;

[0200] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 20; and a light chain variable domain (VL) comprising

[0201] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 21 ;

[0202] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 22;

[0203] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 23.

[0204] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “4D2”.

[0205] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 66; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 54.

[0206] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 66; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 54.

[0207] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “4D2”.

[0208] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 24;

[0209] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 25;

[0210] (iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 26; and a light chain variable domain (VL) comprising

[0211] (i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;

[0212] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 22;

[0213] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 27.

[0214] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0215] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 24, or a functional variant or functional fragment thereof;

[0216] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 25, or a functional variant or functional fragment thereof;

[0217] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 26, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0218] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 21 , or a functional variant or functional fragment thereof;

[0219] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 22, or a functional variant or functional fragment thereof;

[0220] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 27, or a functional variant or functional fragment thereof.

[0221] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0222] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 24;

[0223] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 25;

[0224] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 26; and a light chain variable domain (VL) comprising

[0225] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 21 ;

[0226] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 22; (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 27.

[0227] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “4E6”.

[0228] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 67; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 55.

[0229] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 67; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 55.

[0230] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “4E6”.

[0231] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0232] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 28;

[0233] (ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 29;

[0234] (iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 30; and a light chain variable domain (VL) comprising

[0235] (i) a CDR-L1 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 31 ;

[0236] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 32;

[0237] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 33.

[0238] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0239] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 28, or a functional variant or functional fragment thereof; (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 29, or a functional variant or functional fragment thereof;

[0240] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 30, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0241] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 31 , or a functional variant or functional fragment thereof;

[0242] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 32, or a functional variant or functional fragment thereof;

[0243] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 33, or a functional variant or functional fragment thereof.

[0244] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0245] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 28;

[0246] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 29;

[0247] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 30; and a light chain variable domain (VL) comprising

[0248] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 31 ;

[0249] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 32;

[0250] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 33.

[0251] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “4F5”.

[0252] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 64; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 60.

[0253] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 64; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 60.

[0254] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “4F5”.

[0255] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0256] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 34;

[0257] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 35;

[0258] (iii) a CDR-H3 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 36; and a light chain variable domain (VL) comprising

[0259] (i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37;

[0260] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 38;

[0261] (iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 39.

[0262] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0263] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 34, or a functional variant or functional fragment thereof;

[0264] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 35, or a functional variant or functional fragment thereof;

[0265] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 36, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0266] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 37, or a functional variant or functional fragment thereof;

[0267] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 38, or a functional variant or functional fragment thereof;

[0268] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 39, or a functional variant or functional fragment thereof.

[0269] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0270] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 34;

[0271] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 35;

[0272] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 36; and a light chain variable domain (VL) comprising

[0273] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 37; (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 38;

[0274] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 39.

[0275] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “1 F5”.

[0276] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 70; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 62.

[0277] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 70; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 62.

[0278] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “1 F5”.

[0279] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0280] (i) a CDR-H1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 40;

[0281] (ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 41 ;

[0282] (iii) a CDR-H3 comprising an amino acid sequence having at least 93% sequence identity to SEQ ID NO: 42; and a light chain variable domain (VL) comprising

[0283] (i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 43;

[0284] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 44;

[0285] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45.

[0286] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0287] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 40, or a functional variant or functional fragment thereof; (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 41 , or a functional variant or functional fragment thereof;

[0288] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 42, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0289] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 43, or a functional variant or functional fragment thereof;

[0290] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 44, or a functional variant or functional fragment thereof;

[0291] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 45, or a functional variant or functional fragment thereof.

[0292] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0293] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 40;

[0294] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 41 ;

[0295] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 42; and a light chain variable domain (VL) comprising

[0296] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 43;

[0297] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 44;

[0298] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 45.

[0299] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “3C3”.

[0300] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 69; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 57.

[0301] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 69; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 57.

[0302] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “3C3”.

[0303] In one example provided herein, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0304] (i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 46;

[0305] (ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 47;

[0306] (iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 48; and a light chain variable domain (VL) comprising

[0307] (i) a CDR-L1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 49;

[0308] (ii) a CDR-L2 comprising an amino acid sequence having at least 66 or 100% sequence identity to SEQ ID NO: 50;

[0309] (iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 51.

[0310] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0311] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 46, or a functional variant or functional fragment thereof;

[0312] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 47, or a functional variant or functional fragment thereof;

[0313] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 48, or a functional variant or functional fragment thereof; and a light chain variable domain (VL) comprising

[0314] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 49, or a functional variant or functional fragment thereof;

[0315] (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 50, or a functional variant or functional fragment thereof;

[0316] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 51 , or a functional variant or functional fragment thereof.

[0317] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising

[0318] (i) a CDR-H1 comprising an amino acid sequence of SEQ ID NO: 46;

[0319] (ii) a CDR-H2 comprising an amino acid sequence of SEQ ID NO: 47;

[0320] (iii) a CDR-H3 comprising an amino acid sequence of SEQ ID NO: 48; and a light chain variable domain (VL) comprising

[0321] (i) a CDR-L1 comprising an amino acid sequence of SEQ ID NO: 49; (ii) a CDR-L2 comprising an amino acid sequence of SEQ ID NO: 50;

[0322] (iii) a CDR-L3 comprising an amino acid sequence of SEQ ID NO: 51 .

[0323] An antibody with a variable domain comprising these specific CDRs is referred to the examples below as “12F6”.

[0324] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 71 ; and a light chain variable domain (VL) having at least 95%, preferably at least 99%, sequence identity to SEQ ID NO: 58.

[0325] In one example, the polypeptide according to the invention comprises an antibody variable domain that comprises: a heavy chain variable domain (VH) comprising an amino acid sequence of SEQ ID NO: 71 ; and a light chain variable domain (VL) comprising an amino acid sequence of SEQ ID NO: 58.

[0326] An antibody with a variable domain comprising these specific VH and VL domains is referred to the examples below as “12F6”.

[0327] The antibody variable domain CDR sequences described above have been isolated from (clones of) anti-C1 q antibody producing B cells of human individuals and were found to specifically bind solid-phase C1 q (see examples below). As such, they are particularly suitable objects of the present invention. The sequences of the polypeptides according to the invention have been obtained by sequencing the (clones of the) isolated B cells. They have been sequenced and analyzed via IMGT V-quest (http: / / www.imgt.org / IMGT_vquest / vquest).

[0328] Thus, preferably, the polypeptide according to the invention is an isolated antibody or antigenbinding fragment thereof.

[0329] In the context of sequences, “identical”, “percent identical”, “sequence identity” or “percent sequence identity” in the context of two or more nucleic acid or amino acid sequences or peptides, refers to two or more nucleic acid or amino acid sequences or peptides that are the same or have a specified percentage of amino acid residues or nucleotides that are the same, when compared and aligned for maximum correspondence over a comparison window, as measured using sequence comparison or sequence alignment. A percentage identity with regard to amino acid sequences or peptides where aligned amino acid positions are not identical is often based on conservative alterations, where particular substitutions are expected to produce a functionally equivalent peptide. The percentage identity can be adjusted upwards to correct for conservative alterations and methods for such adjusting are well-known in the art. For clarity, the percentage identity of a sequence; the “target sequence”, for example a sequence listed herein as a SEQ ID NO, is compared and aligned for maximum correspondence over a comparison window, as measured using sequence comparison or sequence alignment, wherein the length of the individually specified contiguous sequence of the comparison window of the target sequence, for example a sequence listed herein as a SEQ ID NO, is the same or substantially the same as the total length of the target sequence.

[0330] A percentage identity between any two nucleic acid sequences, amino acid sequences or peptides can be determined via sequence comparison or sequence alignment. Methods of sequence comparison and sequence alignment are well-known in the art and can be determined via manual alignment and visual inspection or an algorithm, which is suitably implemented on a computer. When performing sequence comparison or alignment one sequence is typically used as a reference sequence to which the other sequence is compared. The comparison occurs in a comparison window which is an individually specified contiguous sequence of each of the compared sequences. Additions or deletions relative from one sequence to the other may be included in any of the sequences, thereby introducing so-called gaps in the other sequence. The introduction of gaps can result in a better alignment between the two sequences. However, the number of gaps in an alignment should be kept to a minimum in order to create a useful alignment, because too many gaps can cause an alignment to become meaningless. To avoid a high sequence identity between two sequences because of the introduction of too many gaps it is known to a person skilled in the art to use a gap penalty in order to compensate. Gap penalties are used to adjust alignment scores based on the number and length of gaps. Examples of gap penalties are constant, linear, affine, convex, and profile-based gap penalties.

[0331] Functional variants of the sequences of the SEQ ID NOs described herein are also encompassed by the embodiments of the present invention. Preferably, a polypeptide according to the invention comprises a functional variant. Functional variants are understood herein to refer to polypeptides having substantially similar amino acid sequences that are biologically active, in that they retain the biological activity of the corresponding SEQ ID NO described herein (i.e., they have substantially similar amino acid sequences to a SEQ ID NO described herein and retain the specific binding to human solid-phase complement component C1q described herein for the corresponding SEQ ID NO). In this example, the substantially similar amino acid sequence may be a sequence which varies from the corresponding SEQ ID NO by (one or more) conservative amino acid substitutions only.

[0332] Functional variants as described herein include polypeptides comprising at least one amino acid sequence of the SEQ ID NOs described herein, wherein the amino acid sequence is changed by substitution, insertion, deletion or addition of at least one amino acid on at least one position in a sequence of a SEQ ID NO. Substitution, insertion or addition of at least one amino acid on at least one position in a sequence of a SEQ ID NO may comprise the substitution, insertion or addition by a non-proteinogenic amino acid. Non-proteinogenic amino acids herein are understood to mean amino acids that are not naturally encoded or found in the genetic code of any organism and are not translationally incorporated into proteins. They comprise any organic compound with an amine and a carboxylic acid functional group. Non-limitative examples of these are phenylalanine derivatives such as 4-methyl-phenylalanine and 3,4-dihydroxy-phenylalanine; phenylglycine derivatives such as 4- hydroxy-phenylglycine; tryptophan derivatives such as 6-amino-7-hydroxy-l-tryptophan; methionine derivatives such as nitrilo-l-methionine; alanine derivatives such as adamanthane; cysteine derivatives such as penicillamine; asparagine / glutamine derivatives such as cysteine-s-acetamide; lysine derivatives such as 2,3-diaminopropanoic acid; arginine derivatives such as c-gamma-hydroxy arginine; serine / threonine derivatives such as homoserine and phosphothreonine; histidine derivatives such as 2-fluoro-l-histidine and asparagine / glutamine derivatives such as l-2-amino-6-methylene- pimelic acid and 4-fluoro-glutamic acid. More examples are for example illustrated in the SwissSidechain database (http: / / www.swisssidechain.ch) (Gfeller et al., Nucleic Acids Research, 2013).

[0333] Substantially similar amino acid sequences have only a small number of sequence changes, for example in non-conserved residues. Functional variants thus preferably comprise variants that have sequence changes that do not affect function, for example in non-conserved residues. Preferably, functional variants have an amino acid sequence comprising at least 66, 85, 87, 88, 90, 92, 93, 94, 95 or 96% sequence identity to any of the amino acid sequences of the SEQ ID NOs described herein, wherein the percentage sequence identity of an amino acid sequence of a functional variant to a respective amino acid sequence of a SEQ ID NO is such that only 1 amino acid is changed in the amino acid sequence of the functional variant with respect to the respective amino acid sequence of the SEQ ID NO. Such a percentage of sequence identity, where only 1 amino acid is changed in a respective amino acid sequence, is expected to not substantially alter the biological function of the polypeptides according to the invention. However, functional variants may also comprise variants having an amino acid sequence that has more than 1 amino acid changed with regard to a respective SEQ ID NO.

[0334] Changes in a nucleic acid sequence by mutation, substitution, insertion, deletion or addition that lead to changes in the amino acid sequence of the encoded peptide, but without altering its biological activity (i.e., specific binding to human solid-phase complement component C1q), are well- known in the art. These are also known as conservative alterations. For example, a codon for alanine, a hydrophobic amino acid, may be substituted by a codon encoding another less hydrophobic amino acid, such as glycine, or a more hydrophobic amino acid, such as valine, leucine, or isoleucine. Similarly, changes which result in substitution of one negatively charged amino acid for another, such as aspartic acid for glutamic acid, or one positively charged amino acid for another, such as lysine for arginine, can also be expected to produce a functionally equivalent product. Nucleotide changes at the N-terminal and / or C-terminal ends of a non-cyclic peptide are also expected to not alter the biological activity of the non-cyclic peptide.

[0335] Preferably, a functional variant has the same or an increased affinity for solid-phase C1 q and the same or a reduced affinity for fluid-phase C1 q, both relative to that of a corresponding unchanged polypeptide according to the invention. A conjugate comprising a polypeptide according to the invention (i.e., a polypeptide comprising an antibody variable domain as described herein) conjugated to or recombinantly fused to a diagnostic agent, detectable agent, or therapeutic agent is also provided herein.

[0336] In one example, the conjugate comprises a linker, for example between the antibody variable domain described herein and the diagnostic agent, detectable agent, or therapeutic agent of the conjugate. Linkers can for example improve folding and stability of the conjugate, expression of the conjugate, bioactivity of the conjugate, pharmacodynamics or pharmacokinetics of the conjugate, or target the conjugate to specific locations in vivo.

[0337] A polypeptide according to the invention can be produced recombinantly. Such a recombinant polypeptide can be recombinantly fused to a diagnostic agent, detectable agent, or therapeutic agent, by for example introducing a nucleotide sequence encoding a diagnostic agent, detectable agent, or therapeutic agent, behind, in front or in the nucleotide sequence of the nucleotide sequence encoding the polypeptide according to the invention. A person skilled in the art understands that such an introduction would retain the intended purpose and effect of the polypeptide according to the invention of being a specific solid-phase C1q binding polypeptide.

[0338] A nucleotide sequence encoding a diagnostic agent, detectable agent, or therapeutic agent may comprise a linker sequence encoding a linker, thereby recombinantly fusing the polypeptide according to the invention to the agent via the linker. A linker may be an amino acid sequence, a peptide, or a polypeptide and may be linear or circular. A linker or linker sequence may be designed such that the agent remains attached in order to couple the location of the polypeptide of the invention to the diagnostic agent, detectable agent, or therapeutic agent. Such a linker or linker sequence is preferably resistant or invulnerable to protease activity from proteases present in the organism to which the conjugate will be administered or subjected. Alternatively, when required such as for example when a therapeutic agent is to be released near the binding location of a polypeptide according to the invention, a linker or linker sequence may be such that it is disrupted by for example protease activity or other means, enabling release of the agent.

[0339] Preferably, the linker is flexible and / or has a sufficient length, in order to sufficiently spatially separate the polypeptide according to the invention from the diagnostic agent, detectable agent, or therapeutic agent while allowing either or both the polypeptide according to invention and the agent to change conformation in accordance with their function. As such, a flexible linker allows for mobility of the connecting domains. Preferably, the linker sequence is rigid and / or has a sufficient length, in order to sufficiently spatially separate the polypeptide according to the invention from the diagnostic agent, detectable agent, or therapeutic agent while allowing either or both the polypeptide according to invention and the agent to change conformation in accordance with their function. As such, a rigid linker allows to keep a fixed distance between the connecting domains.

[0340] Various linkers are known in the art and can be used for the present invention. Examples of linkers are described in Fusion Protein Linkers: Property, Design and Functionality, Chen et al., 2013. A diagnostic agent is a substance that can be used to aid in the diagnosis or monitoring of a disease. It can be administered in vivo or to a subject and it can be used to determine the location or status of a disease causing process.

[0341] A detectable agent is a substance that can be used to determine the presence of a desired molecule, such as a polypeptide according to the invention, in a sample, in vivo, or in a subject.

[0342] Many different diagnostic or detectable agents are known in the art, examples of which are inorganic or organic compounds, radioactive tracers, dyes, proteins, and peptides. Tracers in general and other tracers such as stable isotopes or radioisotopes are also examples of diagnostic or detectable agents. Preferably, a diagnostic or detectable agent is selected from the group consisting of a radionuclide, a radiological contrast agent, a paramagnetic ion, a metal, a fluorescent label, a chemiluminescent label, an ultrasound contrast agent, and a photoactive agent. Preferably, a diagnostic or detectable agent is selected from the group consisting of a radionuclide, a radiological contrast agent, a paramagnetic ion, a metal, a label, for example a fluorescent label or a chemiluminescent label, an ultrasound contrast agent, and a photoactive agent. Many different labels are known in the art, and a label typically comprises or consists of a detectable substance which is attached to the molecule to be detected.

[0343] A therapeutic agent is a substance used to treat, cure, prevent, or ameliorate a disease or to promote well-being. Many different therapeutic agents are known in the art, examples of which are small molecules, oligonucleotides, radionuclides, prodrugs, cytotoxic or cytostatic agents such as chemotherapeutic agents or toxins, photoactive agents, anti-angiogenic agents, enzymes, immunomodulators, cytokines, growth factors, chemokines, hormones, or recombinant proteins. A specific example is a bi-, or multi-specific antibody, wherein at least one arm comprises a polypeptide according to the invention and another arm comprises an agent binding an antigen of interest (wherein the antigen of interest is not solid-phase C1q).

[0344] A polypeptide according to the invention can also be conjugated to a diagnostic agent, detectable agent, or therapeutic agent. Various methods and techniques are known in the art to conjugate various agents to polypeptides, which can also be used for producing the polypeptide according to the invention conjugated to a diagnostic agent, detectable agent, or therapeutic agent. Conjugation can be done post-translationally by chemical crosslinking of the polypeptide according to the invention to the agent by using crosslinking reagents, thereby generating a covalent or a non- covalent bond between the polypeptide and the agent. Crosslinking reagents preferably have similar functions as linkers.

[0345] Various crosslinking reagents and methods are known in the art and can be used for the present invention. Examples of such reagents and techniques are described in Chemical Crosslinking: Role in Protein and Peptide Science, Arora et al., 2017.

[0346] By recombinantly fusing or conjugating a diagnostic agent, detectable agent, or therapeutic agent to the polypeptide according to the invention, the polypeptide can be used in vivo to target the locations where C1 q is deposited (solid-phase), while the polypeptide is not inhibited by the presence of circulating (fluid-phase) C1q. This enables efficient pharmacodynamics by directing a diagnostic agent, detectable agent, or therapeutic agent, such as for example active compounds, drugs, regulators, ortracers, specifically to the locations where C1q-mediated complement activation is taking place. As such, the polypeptide according to the invention can be used as a specific in vivo tracing tool to visualize the locations where C1q is deposited (solid-phase) indicating classical pathway mediated organ damage. This in turn enables targeting of solid-phase C1 q for diagnostic, prognostic, or therapeutic purposes.

[0347] In a particular example, in the context of a polypeptide or conjugate (comprising a polypeptide of the invention), the polypeptide is an antibody, such as a human antibody or a humanized antibody.

[0348] The polypeptides or conjugates according to the invention can be useful for applications in humans. Therefore, a non-human antibody increases the risk of the immune system recognizing the polypeptide or conjugate as foreign and mounting an immune response (i.e., the antibody is immunogenic), and this can be undesirable. In this instance, humanized antibodies can be used. Humanized antibodies are antibodies from non-human species whose protein sequences have been modified to increase their similarity to antibody variants produced naturally in humans, thereby reducing their immunogenicity.

[0349] In one example, the polypeptide according to the invention is selected from the group consisting of a: whole antibody, bispecific antibody, diabody, triabody, tetrabody, minibody, nanobody, sdAb, scFv, di-scFv, bis-scFv, tri-scFv, scFv-Fc, Fab, Fab', F(ab')2, and Fv. For the avoidance of doubt, each of these polypeptides comprise at least one antibody variable domain as described elsewhere herein.

[0350] The aforementioned antibodies are examples of antibodies or antibody fragments produced for example using recombinant antibody techniques and are well-known in the art. As would be clear to a person of skill in the art, some of these formats include two (or more) antibody variable domains. In these formats, at least one of the antibody variable domains specifically binds to human solid-phase C1 q - the additional antibody variable domains may also specifically bind to human solid-phase C1q (e.g., whole antibody) or may specifically bind to an antigen that is not human solid-phase C1 q (e.g., bispecific antibody).

[0351] In one example, the polypeptide according to the invention is a monoclonal antibody. A monoclonal antibody is an antibody produced from a unique leukocyte or a clone of the unique leukocyte. Hereby, the same antibody can be produced from a population of cloned leukocytes.

[0352] In one example, the polypeptide according to the invention further comprises (in addition to the antibody variable region) a heavy chain constant region, optionally wherein the heavy chain constant region is selected from the group consisting of: IgG, IgM, IgE, IgD and IgA, further optionally wherein the heavy chain constant region is selected from the group consisting of: lgA1 , lgA2, lgG1 , lgG2, lgG3 and lgG4.

[0353] Human antibodies are classified into five isotypes or classes according to their heavy chain constant regions: IgG, IgM, IgE, IgD and IgA. IgA and IgG have respectively 2 and 4 subclasses. The classes differ in their biological properties and can therefore be used to adapt the effects of the polypeptide according to the invention. For example, IgA can prevent colonization by pathogens, while IgD can activate basophils and mast cells to produce antimicrobial factors.

[0354] In one example, the polypeptide according to the invention comprises a modified Fc domain, for example wherein the Fc domain is modified to increase, decrease, or prevent interaction with a neonatal Fc receptor (FcRn), to increase, decrease, or prevent hexamerisation, to increase, decrease, or prevent interaction with a Fc receptor (FcR), or to increase, decrease, or prevent binding of C1 q via the Fc domain of the polypeptide. An increase, decrease, or prevention is chosen based on the intended function of the polypeptide according to the invention. For example, hexamerisation of IgG is proposed to be crucial for efficient interaction with C1 q and subsequent complement activation. Van Osch et al. (Fc Galactosylation Promotes Hexamerization of Human lgG1 , Leading to Enhanced Classical Complement Activation, 2021) show that various mutations can increase galactolysation of a conserved N-linked glycan in the Fc region of human lgG1 and enhance lgG1 hexamerisation, thereby increasing complement activation. Modifying a polypeptide according to the invention as such could be useful for using the polypeptide for the treatment of cancer or bacterial infections. Alternatively, modifying the polypeptide such that hexamerisation is decreased could be useful for using the polypeptide for the treatment of auto-immune disease. FcR and FcRn are expressed on immune cells and endothelial cells, involved in recognizing antibody Fc domains and subsequent clearance of complexes recognized by the antibodies. Modifying the Fc domains of the polypeptides according to the invention can therefore increase or decrease clearance of targets recognized by solidphase C1q.

[0355] The Fc domain may be modified to be for example a Fc silent or Fc active variant, by introducing mutations in the nucleotide sequences encoding the Fc regions. A Fc silent variant loses the ability to trigger additional complement activation and / or Fc-receptors, while a Fc active variant acquires the capacity to enhance complement activation and / or Fc-receptors. Similarly, the Fc domain may be modified to have an increased or reduced interaction (e.g. binding or binding affinity) with an Fc receptor compared to an unmodified Fc domain. Preferably, the unmodified Fc domain has been modified and this unmodified Fc domain is used to compare the resulting increased or reduced interaction (e.g. binding or binding affinity) with the Fc receptor.

[0356] Hence, another aspect of the invention is a polypeptide or conjugate according to the invention, wherein the modified Fc domain has increased or reduced interaction with an Fc receptor compared to an unmodified Fc domain of the polypeptide.

[0357] The Fc domain may be modified by changing the nucleic acid sequence encoding the Fc domain by mutation, substitution, insertion, deletion, or addition of one or more nucleic acids of the sequence. Alternatively, the Fc domain may be modified by post-translational chemical modification. The Fc domain may be modified by biochemical or enzymatic modification. The Fc domain may be modified by a modification to a glycan structure, a glycosylation site, or number of glycosylation sites. Various means to accomplish these modifications are known in the art, see for example K.O. Saunders: “Conceptual Approaches to Modulating Antibody Effector Functions and Circulation Half- Life”, Front Immunol., 7 June 2019, doi: 10.3389 / fimmu.2019.01296, R. Liu et al.: “Fc-Engineering for Modulated Effector Functions — Improving Antibodies for Cancer Treatment”, Antibodies (Basel), 2020 Dec; 9(4): 64, and B. Ma et al.: “Protein Glycoengineering: An Approach for Improving Protein Properties”, Front. Chem., 23 July 2020, doi: 10.3389 / fchem.2020.00622.

[0358] Hence, another aspect of the invention is a polypeptide or conjugate according to the invention, wherein the modified Fc domain comprises an amino acid mutation, substitution, insertion, deletion, or addition, a modification to a glycan structure, a glycosylation site, or number of glycosylation sites, or a post-translational chemical, biochemical, or enzymatic modification. Preferably, the modification to a glycan structure, a glycosylation site, or number of glycosylation sites comprises afucosylation, preferably wherein the afucosylation is at amino acid residue N297 of a heavy chain constant region. Preferably, the modification to a glycan structure, a glycosylation site, or number of glycosylation sites comprises a biantennary glycan, preferably wherein the biantennary glycan is at amino acid residue N297 of a heavy chain constant region.

[0359] Another aspect of the invention is a polypeptide or conjugate according to the invention, wherein the polypeptide further comprises a heavy chain constant region comprising one or more amino acid substitutions. Preferably, the one or more amino acid substitutions comprises S298A, E333A, and K334A. Preferably, the one or more amino acid substitutions comprises S239D and I332E. Preferably, the one or more amino acid substitutions comprises S239D, A330L, and I332E. Preferably, the one or more amino acid substitutions comprises G236A. Preferably, the one or more amino acid substitutions comprises G236A, S239D, and I332E. Preferably, the one or more amino acid substitutions comprises G236A, A330L, and I332E. Preferably, the one or more amino acid substitutions comprises G236A, S239D, A330L, and I332E. Preferably, the one or more amino acid substitutions comprises F243L, R292P, Y300L, V305I, and P396L. Preferably, the one or more amino acid substitutions comprises L235V, F243L, R292P, Y300L, and P396L. Preferably, the one or more amino acid substitutions comprises P247I and A339Q. Preferably, the one or more amino acid substitutions comprises L234Y, G236W and S298A.

[0360] Preferably, the one or more amino acid substitutions comprises L234F, L235E, and P331S. Preferably, the one or more amino acid substitutions comprises N298A. Preferably, the one or more amino acid substitutions comprises K326W and E333S. Preferably, the one or more amino acid substitutions comprises S267E, H268F, and S324T. Preferably, the one or more amino acid substitutions comprises L235E. Preferably, the one or more amino acid substitutions comprises L234A and L235A. Preferably, the one or more amino acid substitutions comprises L234A, L235A, and P329G. Preferably, the one or more amino acid substitutions comprises D265A. Preferably, the one or more amino acid substitutions comprises G237A. Preferably, the one or more amino acid substitutions comprises E318A. Preferably, the one or more amino acid substitutions comprises E233P. Preferably, the one or more amino acid substitutions comprises G236R and L328R. Preferably, the one or more amino acid substitutions comprises L234A, L235A, G237A, P238S, H268A, A330S, and P331S. Preferably, the one or more amino acid substitutions comprises N297A, N297G, or N297Q.

[0361] The numbers above indicate the amino acid residue numbers of the heavy chain constant region. The above amino acid substitutions are examples of a modified Fc domain according to the invention. Preferably, the heavy chain constant region is selected from the group consisting of: IgG, IgM, IgE, IgD and IgA. Preferably, the heavy chain constant region is selected from the group consisting of: lgA1 , lgA2, lgG1 , lgG2, lgG3 and lgG4. Preferably, the heavy chain constant region is IgG. Preferably, the heavy chain constant region is lgG2. Preferably, the heavy chain constant region is lgG3. Preferably, the heavy chain constant region is lgG4. More preferably, the heavy chain constant region is lgG1.

[0362] Another aspect of the invention is an isolated nucleic acid comprising a sequence encoding a polypeptide or a conjugate according to the invention. Preferably, the sequence comprises a nucleic acid sequence encoding a modified Fc domain, more preferably the nucleic acid sequence encoding the modified Fc domain comprises a mutation, substitution, insertion, deletion, or addition of one or more nucleic acids of the sequence.

[0363] Deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) are nucleic acids. In polymeric form DNA and messenger RNA encode the genetic information that results in the generation of peptides, polypeptides, and proteins. A nucleic acid according to the invention can therefore be used to produce the polypeptide according to the invention. As such, an isolated nucleic acid encoding a polypeptide or a conjugate according to the invention is also part of the present invention. DNA and RNA can be chemically modified to for example increase or decrease stability or to increase or decrease the rate of expression of the genetic information contained within.

[0364] Another aspect of the invention is an isolated cell comprising a nucleic acid according to the invention.

[0365] A cell is the basic structural and functional unit of life. Every cell comprises a cytoplasm enclosed within a lipid membrane, which contains various molecules such as proteins and nucleic acids. A cell according to the invention can therefore be used to produce the polypeptide according to the invention.

[0366] Another aspect of the invention is a pharmaceutical composition comprising a polypeptide or conjugate according to the invention, and a pharmaceutically acceptable excipient, adjuvant, diluent, or carrier.

[0367] A pharmaceutically acceptable excipient, adjuvant, diluent, or carrier includes any of the standard pharmaceutically acceptable excipient, adjuvant, diluent, or carrier known in the art, such as water, phosphate buffered saline, emulsions such as a water and oil emulsion, and various types of wetting agents.

[0368] A pharmaceutical composition according to the invention may also include more than one active compound for the disease to be treated. Preferably, the pharmaceutical composition comprises a polypeptide according to the invention and at least one additional active compound that do not adversely affect each other.

[0369] Another aspect of the invention is a kit comprising a polypeptide or conjugate according to the invention, and instructions for using the polypeptide or conjugate to (i) treat or prevent auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in a subject in need of such treatment, or (ii) diagnose or predict the development of auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1 q, in vivo in a subject.

[0370] Another aspect of the invention is a kit comprising a polypeptide or conjugate according to the invention, and instructions for using the polypeptide or conjugate to (i) treat or prevent infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in a subject in need of such treatment, or (ii) diagnose or predict the development of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in vivo in a subject.

[0371] The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used as a medicament. By specifically targeting solid-phase C1q, the polypeptide can be used to either inhibit an immune response (by inhibiting activation of the complement cascade by C1q) or enhance or activate an immune response (by enhancing or activating the complement cascade via C1q). In addition, following binding of the polypeptide according to the invention to solid-phase C1q a number of biological mechanisms can be triggered depending on the molecular nature of the Fc domain. Examples can be but are not limited to additional complement activation to allow immunecomplex solubilisation, transport, and degradation, or, in other settings, allowing interaction with cells expressing Fc-receptors now interacting with the polypeptide according to the invention bound to the solid-phase C1q, thereby allowing binding to and / or uptake by phagocytic cells. Other Fc-receptor- mediated functions, for example antibody-dependent cellular cytotoxicity via NK cells, may also be enhanced due to the interaction of the polypeptide according to the invention bound to solid-phase C1 q with the Fc-receptor. As such, a pro-inflammatory response and / or elimination of C1q-opsonized cells or objects may be enhanced. Conversely, modifications in for example the Fc domain of the polypeptides of the present invention (as disclosed above) may inhibit interaction with cells expressing Fc-receptors with the polypeptide according to the invention bound to the solid-phase C1 q, thereby inhibiting binding to and / or uptake by phagocytic cells. Other Fc-receptor-mediated functions, for example antibody-dependent cellular cytotoxicity via NK cells, may similarly be inhibited due to the modified polypeptide according to the invention bound to solid-phase C1q reducing interaction of the Fc-receptor with C1q-opsonized objects. As such, a pro-inflammatory response and / or elimination of C1 q-opsonized cells or objects may be inhibited. The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to treat or prevent infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, in a subject in need of such treatment.

[0372] The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used in immunotherapy or transplantation, preferably wherein the use in transplantation comprises use in the pre-treatment of a subject prior to receiving transplantation of an organ, tissue or cells, use in the pre-treatment of an organ, tissue or cells prior to transplantation to a recipient subject, or use in the treatment or prevention of tissue or organ rejection in a subject.

[0373] The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to treat or prevent auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in a subject in need of such treatment.

[0374] A disease or disorder associated with complement activation involving C1 q may be associated directly with C1q, such as mutations in the C1q protein, or associated with elements that interact directly with C1 q, such as C1 r or C1 s or via interactions with the globular domain or the collagen-like domain of C1q, or associated with elements that interact indirectly with C1q, such as for example further downstream components of the classical complement activation pathway. In addition, a disease or disorder associated with complement activation involving C1q may be associated with a subject’s own antibodies or administered antibodies recognizing antigens in the subject to an extent which either over- or underactivates the complement pathway involving C1q. Such bound antibodies present a major target for C1q. The situation described above can then cause destruction of the subject’s own cells (e.g. autoimmune disease) or tissues or mediate destruction of cells or agents that are detrimental to the health of the subject (e.g. cancer or virus).

[0375] Diseases or disorders that are associated with complement activation are readily identifiable by a person of skill in the art. For example, they may be an autoimmune disease or disorder that involves C1 q, such as but not limited to: SLE, rheumatoid arthritis, or cold agglutinin disease.

[0376] Other diseases or disorders associated with complement activation involving C1 q may be transplantation associated pathology, such as but not limited to hyperacute rejection, chronic rejection, antibody mediated rejection, and ABO-incompatibility, of any of the following: solid organs, bone marrow, hematopoietic stem-cell transplantation and all other cellular products including but not limited to platelets, erythrocytes, leukocytes, stem cells, organoids, and CAR-T cells. Moreover, C1 q is involved in depletion strategies prior to transplantation such as but not limited to anti-thymocyte globulin and anti-CD52 treatment.

[0377] Further diseases or disorders associated with complement activation involving C1 q may be neurological diseases or disorders such as but not limited to Guillain-Barre syndrome (GBS) and Multifocal Motor Neuropathy.

[0378] Also, diseases or disorders associated with complement activation involving C1q may be neurodegenerative diseases or disorders such as but not limited to Alzheimer's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, Down syndrome, Parkinson's disease, and Huntington's disease.

[0379] In addition, diseases or disorders associated with complement activation involving C1 q may comprise the development or progression of cancer. During development or progression of cancer different antigens may be presented by the cancer cells, which may indirectly and / or preferentially recruit C1q to the progressing cancer cell, thereby presenting a target for treatment using the polypeptides according to the invention.

[0380] Diseases or disorders associated with complement activation involving C1q may include ischemia-reperfusion injury. Ischemia-reperfusion injury activates the complement system via release of damage associated molecular patterns from acutely injured tissue, which then enhances a further immune response. Administering polypeptides according to the invention before, during, or directly following surgery causing ischemia-reperfusion injury may therefore treat or prevent the ischemiareperfusion injury.

[0381] Diseases or disorders associated with complement activation involving C1q may also be due to a response of the immune system directed to bacterial infections, myco-bacterial infections, or viral infections. Such a response involving C1q may be overactive or not sufficient, which can both respectively be addressed using the polypeptides according to the invention.

[0382] The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to enhance treatment or prevention by other medicaments, of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, in a subject in need of such treatment. The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to enhance immunotherapy or transplantation by other medicaments, preferably wherein the transplantation comprises the pre-treatment of a subject prior to receiving transplantation of an organ, tissue or cells, the pre-treatment of an organ, tissue or cells prior to transplantation to a recipient subject, orthe treatment of tissue ororgan rejection in a subject, in a subject in need of such treatment. The polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to enhance treatment or prevention by other medicaments, of auto-immune disease, cancer, ora disease or disorder associated with complement activation involving C1 q, in a subject in need of such treatment. As such, a polypeptide, conjugate, or pharmaceutical composition according to the invention can be used as a medicament complementary to other medicaments, preferably without adversely affecting the other medicaments. Suitable treatment or prevention by other medicaments comprises medicaments that intentionally or unintentionally activate, enhance, and / or initiate complement activation involving C1q. Such medicaments are readily identifiable by a person of skill in the art. An aspect of the invention is thus a polypeptide, conjugate, or pharmaceutical composition for use according to the invention, wherein the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more other pharmaceutical compositions, preferably without adversely affecting the other pharmaceutical compositions. Suitable other pharmaceutical compositions comprise pharmaceutical compositions that intentionally or unintentionally activate, enhance, and / or initiate complement activation involving C1q. Such compositions are readily identifiable by a person of skill in the art.

[0383] An example of these other medicaments or pharmaceutical compositions is a medicament or pharmaceutical composition for a treatment for a specific population of cells, which involves administering antibodies targeting the population in order to reduce the population in numbers. The population of cells may be but is not limited to B cells, T cells, natural killer cells, dendritic cells, and neutrophils, and any sub-population of these cells such as but not limited to CD4+ T cells, CD8+ T cells, yb T cells, iNKT cells, and Tregs. All these have been successfully used to selectively deplete immune cells in vivo and can be used for the treatment of cancer or autoimmune disease.

[0384] A specific example of such a treatment is anti-CD20 mediated depletion of B cells, targeting the CD20 positive population of B cells. Anti-CD20 mediated depletion of B cells by commercially available CD20-targeting monoclonal antibodies such as rituximab, ocrelizumab, obinutuzumab, ofatumumab, ocaratuzumab, ibritumomab tiuxetan, tositumomab, veltuzumab, and ublituximab, has proven successful for treating B cell malignancies, such as but not limited to diffuse large B-cell lymphoma, follicular lymphoma, and chronic lymphocytic leukemia, and many autoimmune diseases, such as but not limited to multiple sclerosis, rheumatoid arthritis, pemphigus, pemphigoid, myasthenia gravis, and neuromyelitis optica. C1 q binds to anti-CD20 bound to B cells, thereby triggering complement dependent cytotoxicity (CDC). The polypeptides according to the invention can thus enhance such a treatment.

[0385] A further example of these other medicaments or pharmaceutical compositions is a medicament or pharmaceutical composition for depletion prior to transplantation, such as but not limited to anti-thymocyte globulin and anti-CD52 treatment, where the same mechanism for the polypeptides according to the invention is envisioned as for anti-CD20 mediated depletion of B cells.

[0386] Another example of these other medicaments or pharmaceutical compositions is a medicament or pharmaceutical composition for the treatment or prevention of cancer. This can be any medicament or pharmaceutical composition for the treatment or prevention of cancer, for example but not limited to chemotherapy, oncolytic virus therapy, CAR-T cell therapy and antibody therapy, in which the therapy destroys the cancer cells. Polypeptides according to the invention can enhance the anti-cancer therapy by for example also targeting the cancer cells or targeting cell fragments released by the destroyed cancer cells. Alternatively, the polypeptides according to the invention can enhance the other medicament or pharmaceutical composition being an antibody according to the same mechanism as for anti-CD20 mediated depletion of B cells.

[0387] Another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition for use according to the invention, wherein the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more diagnostic agents, detectable agents or therapeutic agents, preferably without adversely affecting the other diagnostic agents, detectable agents or therapeutic agents. Preferably, when the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more other pharmaceutical compositions or one or more diagnostic agents, detectable agents or therapeutic agents, the one or more other pharmaceutical compositions or one or more diagnostic agents, detectable agents or therapeutic agents is selected from the group consisting of a: complement activating compound, complement inhibiting compound, tracer, label, radioactive compound, toxin, CRISPR-associated protein (Cas), and nucleic acid. Preferably, when the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more other pharmaceutical compositions, the one or more other pharmaceutical compositions is selected from the group consisting of a: complement activating compound, complement inhibiting compound, radioactive compound, toxin, CRISPR-associated protein (Cas), and nucleic acid. Preferably, when the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more diagnostic agents, detectable agents or therapeutic agents, the one or more diagnostic agents, detectable agents or therapeutic agents is selected from the group consisting of a: complement activating compound, complement inhibiting compound, tracer, label, radioactive compound, toxin, CRISPR-associated protein (Cas), and nucleic acid. Preferably, when the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more diagnostic agents or detectable agents, the one or more diagnostic agents or detectable agents is selected from the group consisting of a: tracer, label, radioactive compound, and nucleic acid.

[0388] Preferably, the Cas is selected from the group consisting of: Cas3, Cas9, Cas12a, Cas12b, Cas13, Cas7-11 , CasMINI, and SuperFi-Cas9. Preferably, the nucleic acid is a ribonucleic acid (RNA) ora deoxyribonucleic acid (DNA). Preferably, the RNA is selected from the group consisting of a: small interfering RNA (siRNA), asymmetrical interfering RNA (aiRNA), microRNA (miRNA), Dicer-substrate RNA (dsRNA), small hairpin RNA (shRNA), messenger RNA (mRNA), antisense RNA (asRNA), aptamer, circular RNA (circRNA), self-amplifying RNA (saRNA), catalytic RNA, anti-miRNA, long noncoding RNA (IncRNA), single-guide RNA (sgRNA), and mRNA encoding a Cas. Preferably, the DNA is selected from the group consisting of a: antisense oligonucleotide (ASO), aptamer, episome, and catalytic DNA. A nucleic acid according to the invention preferably comprises an oligonucleotide molecule. A DNA may have a nucleotide sequence encoding an RNA of interest. An RNA may have a nucleotide sequence encoding a peptide, polypeptide, or protein of interest. For example, an RNA may be an mRNA. The mRNA may encode a polypeptide, which may be translated in a cell to produce the encoded polypeptide.

[0389] In one example, the polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to enhance anti-tumor effector mechanisms in a subject in need thereof, for example by enhancing complement activation on the tumor only. This aspect of the invention may be particularly relevant to a subject that is suffering from cancer. Cancer cells comprise mutated genes and therefore express antigens that can be detected by the immune system. The resulting antibodyantigen complexes can serve as natural ligands for C1q, resulting in solid-phase C1q bound to the complexes, and thereby inducing CDC against these cancer cells. The polypeptide, conjugate or pharmaceutical composition described herein that is specific for solid-phase C1 q can be used to further promote CDC by presenting a natural ligand for additional C1 q to bind, while simultaneously not hindering the attachment of other components of the classical complement pathway to solid-phase C1 q. CDC is thus increased, thereby treating the cancer. Alternatively, or in addition, the polypeptide, conjugate or pharmaceutical composition described herein that is specific for solid-phase C1q, when bound to the solid-phase C1 q bound to the complexes, may be used to promote antibody-dependent cellular cytotoxicity or antibody-dependent cellular phagocytosis via Fc receptors on the surface of immune cells, thereby treating the cancer.

[0390] In another example, the polypeptides, conjugates, and pharmaceutical compositions provided herein may be used to inhibit the complement pathway in a subject in need thereof, for example in a subject suffering from auto-immune disease. Autoantibodies are a hallmark of most auto-immune diseases, which can serve as natural ligands for C1 q, resulting in solid-phase C1q bound to the autoantibodies, and thereby inducing complement dependent cytotoxicity (CDC) against cells expressing antigens that are recognized by the autoantibodies. The polypeptide, conjugate or pharmaceutical composition described herein can be used to specifically bind solid-phase C1q and, for example, due to steric hindrance, can inhibit binding of other components of the classical complement pathway to solid-phase C1q. Complement activation is prevented and organ damage limited. As a consequence of complement inhibition also complement mediated inflammation and CDC is thus inhibited, thereby treating the auto-immune disease. Alternatively, or in addition, the polypeptide, conjugate or pharmaceutical composition described herein that is specific for solid-phase C1 q, when bound to the solid-phase C1q bound to the complexes, may be used to inhibit antibodydependent cellular cytotoxicity or antibody-dependent cellular phagocytosis via Fc receptors on the surface of immune cells, thereby treating the auto-immune disease. Modifications in the Fc domain of the polypeptides according to the invention that prevent interaction of the polypeptides with Fc receptors may be preferred in this context.

[0391] The two examples above may be extended to other diseases having a similar mechanism of action with regard to involvement of C1q, for example infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, and rejection of cells, tissues or organs after transplantation (i.e. via allogeneic immune response).

[0392] Transplantation herein refers to the removal of cells, tissues, or organs from the body of a subject and then their transfer back into the same body (autotransplantation) or into a different body (allotransplantation or xenotransplantation). In the context of the present invention, transplantation preferably refers to allotransplantation or xenotransplantation, in particular to allotransplantation. T ransplantation typically involves surgery, the use of immunosuppressants, the possibility of infection, and / or the possibility of rejection of the transplant.

[0393] Complement activation contributes to allograft injury, for example during ischemia / reperfusion injury and antibody mediated rejection. In addition, the complement system modulates the responses of T cells and B cells to antigens. As such, the polypeptides of the present invention are suitable for use in the treatment and prevention of transplant rejection or graft versus host disease using a similar rationale as for the treatment and prevention of auto-immune disease as discussed herein. Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use as a medicament or a diagnostic.

[0394] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in the treatment of a disease or disorder associated with complement activation involving C1q.

[0395] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in the treatment of auto-immune disease or cancer. In one example, the auto-immune disease or cancer comprises cells expressing an antigen to which C1q directly or indirectly binds, turning it into solid-phase C1 q.

[0396] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in the treatment of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease. In one example, the infectious disease, neurological disease, neurodegenerative disease, graft versus host disease comprises cells expressing an antigen to which C1 q directly or indirectly binds, turning it into solid-phase C1q.

[0397] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in immunotherapy or transplantation, preferably wherein the use in transplantation comprises use in the pre-treatment of a subject prior to receiving transplantation of an organ, tissue or cells, use in the pre-treatment of an organ, tissue or cells prior to transplantation to a recipient subject, or use in the treatment or prevention of tissue or organ rejection in a subject. In one example, the transplanted organ, tissue or cells comprise cells expressing an antigen to which C1 q directly or indirectly binds, turning it into solid-phase C1 q.

[0398] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in a method of diagnosis or prediction of development of autoimmune disease, cancer, or a disease or disorder associated with complement activation involving C1 q, in vivo in a subject.

[0399] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in a method of diagnosis or prediction of development of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, in vivo in a subject.

[0400] Yet another aspect of the invention is a polypeptide, conjugate, or pharmaceutical composition according to the invention for use in the treatment of systemic lupus erythematosus (SLE). It is disclosed herein, in particularwith reference to Figure 14, that the specific polypeptides of the invention may be used for treatment of SLE as they compete for the same epitope on solid-phase C1 q.

[0401] Another aspect of the invention is a polypeptide or conjugate according to the invention for detecting solid-phase C1q. The invention also provides a method of detecting solid-phase C1q in a biological sample, comprising the steps of: a. contacting a polypeptide or conjugate according to the invention with the biological sample; and b. detecting the presence or absence of the polypeptide bound to solid-phase C1 q in the biological sample.

[0402] In one example, the method of detecting solid-phase C1q in a biological sample is an in vitro method.

[0403] The biological sample may be any suitable sample, for example it may be a blood sample, plasma, serum, cerebrospinal fluid, lung aspirate or a homogenized tissue sample.

[0404] Means for contacting and detecting the presence or absence of the polypeptide bound to solidphase C1q are well known in the art, and are described elsewhere herein, for example an ELISA assay may be used.

[0405] The invention also provides a method of detecting solid-phase C1q in vivo in a subject, comprising the steps of: a. administering a polypeptide, conjugate, or pharmaceutical composition according to the invention, to the subject; and b. detecting the presence or absence (or location) of the polypeptide bound to solid-phase C1q in the subject.

[0406] Means for administering a polypeptide, conjugate, or pharmaceutical composition are well known in the art. For example, a polypeptide, conjugate, or pharmaceutical composition may be administered by intravenous infusion, injection, intravitreous application, or oral administration.

[0407] Means for detecting the presence, absence or location of the polypeptide bound to solid-phase C1 q in the subject are also well known. Routine detection means are for example discussed elsewhere herein.

[0408] The invention also provides a method of removing C1 q in vivo in a subject, comprising the steps of: a. administering a polypeptide, conjugate, or pharmaceutical composition according to the invention, to the subject; and b. removing solid-phase C1q or complexes comprising solid-phase C1q from a target tissue to which they are bound. Means for removing solid-phase C1 q or complexes comprising solid-phase C1 q from a target tissue to which they are bound are well known.

[0409] The invention further provides a method of treating a subject, comprising administering a therapeutically effective amount of a pharmaceutical composition according to the invention to a subject in need thereof. A therapeutically effective amount refers to an amount sufficient to reduce the severity and / or duration of a disease or a symptom thereof. Progression, development, or onset of the disease may thereby be reduced or prevented. The amount of polypeptide or conjugate actually administered will typically be determined by a physician, in the light of the relevant circumstances, including the condition to be treated, the chosen route of administration, the actual compound administered, the age, weight, and response of the individual patient, the severity of the patient's symptoms, and the like.

[0410] Sequences and antibody affinities

[0411]

[0412] Table 1 . Sequences

[0413] Table 2. Results of affinity measurements by surface plasmon resonance on Biacore T200. KD (M) indicates the dissociation constant. Ka (1 / Ms) indicates the on-rate. Kd (1 / s) indicates the off-rate.

[0414]

[0415] Table 3. Genetic characteristics of human anti-C1q mAbs as analyzed by IMGT V-quest. CDR3, Complementarity-Determining Region 3; AA, amino acid.

[0416]

[0417]

[0418] Table 4. Variable chain amino acid sequences.

[0419]

[0420] Table 5. Exemplary full variable light chain and variable heavy chain DNA sequences - NB including some ancillary sequence.

[0421] Table 6. Light chain and heavy chain DNA sequences recognized as variable domain by IMGT V- quest.

[0422] Detailed Description of the Figures The invention will now be discussed with reference to the figures, which show preferred exemplary embodiments of the subject invention. Reference is made to the Materials and Methods and Results sections of the Examples for further details on the experimental results as presented in the Figures.

[0423] Figure 1 shows binding of anti-C1q monoclonal antibodies. Different concentrations of the human anti-C1q monoclonal antibodies designated 2D2, 1 F4, 4C11 , 4D2, 4E6, 4F5, 1 F5, 3C3, and 12F6 were assessed for binding to solid-phase C1 q (Figure 1A) in ELISA assays. Figure 1A: concentration of the antibody in ng / ml is shown on the x-axis. Absorption at A=415nm is shown on the y-axis and is a measure for the level of binding of the antibody to solid-phase C1 q. Figure 1A shows binding of the anti-C1q monoclonal antibodies to solid-phase C1q, wherein C1q has been directly coated on the substrate of the wells-plate. Figure 1 B: overview of binding to well-known ligands of polyreactive mAbs. To exclude the possibility that the anti-C1 q monoclonal antibodies are polyreactive, binding to several well-known targets of (polyreactive) autoantibodies was tested in ELISA. For all anti-C1q mAbs, no meaningful binding to single-stranded DNA, lipopolysaccharide (LPS) or insulin was observed, demonstrating specificity of these mAbs to C1q. In addition, a negative control recombinant anti-dinitrophenol (DNP) monoclonal antibody shows no binding to solid-phase C1 q in figure 1 A and no binding to ligands of polyreactive mAbs in figure 1 B. Thus, all antibodies bind to solid-phase C1q to a varying degree depending on their concentration.

[0424] Figures 2 and 3 show binding of anti-C1q monoclonal antibodies to solid-phase C1 q in the presence of fluid-phase C1q as competitor. Different concentrations of the human anti-C1q monoclonal antibodies designated 2D2, 1 F4, 4C11 , 4D2, 4E6, 4F5, 1 F5, 3C3, and 12F6, as well as a negative control mouse anti-C1q monoclonal antibody 4A4B11 , were assessed for binding to solidphase C1 q in ELISA competition assays. Concentration of the antibody in ng / ml is shown on the x- axis. OD at 415nm is shown on the y-axis and is a measure for the level of binding of the antibody to solid-phase C1 q. Solid-phase C1 q was first deposited, after which different concentrations of the antibodies together with or without fluid-phase C1 q as competitor were added. Fluid-phase C1q was either added as purified C1q at 20 pg / ml, or as C1q in 5% or 25% Normal Human Serum (NHS). Next to fluid-phase C1q, NHS contains a multitude of other proteins and therefore acts as a non-specific sink for antibodies. Additions of all three fluid-phase C1 q competitors resulted in decreased binding of the negative control 4A4B11 antibody to solid-phase C1q, when compared to when no competitor was added. On the other hand, binding to solid-phase C1 q of all human anti-C1 q antibodies was not inhibited or only marginally inhibited by addition of fluid-phase C1q competitors. Therefore, all tested human anti-C1q monoclonal antibodies specifically bind to solid-phase C1 q and do not substantially bind to fluid-phase C1q.

[0425] Figure 4 shows binding of anti-C1q monoclonal antibodies to solid-phase C1 q in the presence of fluid-phase C1 q as competitor as a selection of the data presented in figures 2 and 3. Based on each of the titration curves of figures 2 and 3 a linear part was selected for each antibody. A corresponding exemplary concentration of this linear part was used to determine the percentage inhibition by fluid-phase C1q of binding of the antibody to solid-phase C1q. In Figure 4, the “no competition” condition is considered as 100% antibody binding and 0% inhibition. The respective concentrations are considered to give a good indication of possible inhibition as there is sufficient binding of the anti-C1q antibodies to solid-phase C1q. Percentage binding is shown for 0.16 pg / ml (1 F5, 4F5, mouse 4A4B11), 0.64 pg / ml (3C3, 4D2, 4E6), 2.56 pg / ml (1 F4, 2D2, 4C11) and 10.24 pg / ml (12F6) anti-C1q antibody. While for the mouse anti-human C1q mAb 4A4B11 it is clear that all three fluid-phase competitors result in decreased binding of the antibody to solid-phase C1q, it can be seen that all tested human anti-C1q monoclonal antibodies do not display an inhibition, thus specifically bind to solid-phase C1 q and do not substantially bind to fluid-phase C1 q.

[0426] Figure 5 shows competition with fluid-phase purified C1 and C1q depleted serum for selected anti-C1 q mAbs 1 F5, 4F5 and mouse 4A4B11. The full C1 qr2s2 complex (denoted C1) was used as fluid-phase purified C1 competitor. Figure 5 shows the results of similar competition experiments as presented in Figures 2-4.

[0427] Figure 6 shows that binding of anti-C1 q antibodies is inhibited by solid-phase C1 q but not fluidphase C1q. Binding of human 0.1 pg / well anti-C1q antibodies to C1 q coated ELISA was analysed in the presence of 1 pg / well C1 q or Human Serum Albumin (HSA), either in solid-phase or in fluid-phase, as competitor. Absorption at A=415nm is shown on the y-axis and is a measure for the level of binding of the antibody to the coated solid-phase C1 q. Of these competitors, only solid-phase C1q was able to significantly compete for binding by anti-C1 q antibodies, confirming specificity for solid-phase C1 q. Anti-C1 q antibodies were engineered to have inactive Fc and have only one Fab specific for C1 q, thus becoming bispecific. Statistical analysis was performed with one-way ANOVA with Dunnett multiple comparisons test. Asterisks indicate a significant difference, ‘ns’ indicates a non-significant difference.

[0428] Figure 7A shows binding of anti-C1 q monoclonal antibodies to solid-phase C1 q. Experimental conditions are a similar to those of Figure 1A, with the exception that C1 q has now been bound to different natural ligands instead of directly to the ELISA plate. The left graph of Figure 7A shows binding of the anti-C1q monoclonal antibodies to solid-phase C1 q, wherein C1q has been bound to its natural ligand IgG. The middle graph of Figure 7A shows binding of the anti-C1q monoclonal antibodies to solid-phase C1q, wherein C1q has been bound to its natural ligand IgM. The right graph of Figure 7A shows binding of the anti-C1q monoclonal antibodies to solid-phase C1 q, wherein C1q has been bound to its natural ligand C-reactive protein (CRP).

[0429] Figure 7B shows binding of anti-C1q mAbs to IgG opsonized cells as detected by flow cytometry. Geometric mean fluorescence intensity (gMFI) of streptavidin-AlexaFluor647 bound to biotinylated anti-C1q mAbs bound to cells was used as a readout. Biotinylated anti-DNP antibody was used as a negative control. Binding of the anti-C1q mAb was specifically observed to alemtuzumab- opsonized cells in the presence, but not in the absence, of C1 q. Figure 7C shows binding of anti-C1 q mAbs to necrotic cells as detected by flow cytometry. Geometric mean fluorescence intensity (gMFI) of streptavidin-AlexaFluor647 bound to biotinylated anti-C1q mAbs bound to cells was used as a readout. Biotinylated anti-DNP antibody was used as a negative control. Antibody independent physiological C1 q binding to dead cells was analyzed using necrotic PBMCs. C1 q binding on necrotic cells was more modest as detected by the positive control polyclonal antibody. Nevertheless, several anti-C1 q mAbs showed binding to C1q on necrotic cells, especially those identified in previous ELISA experiments as the strongest binding mAbs.

[0430] Figure 8 shows Western blots of C1q in various states of protein folding, with detection by antiCi q mAbs. Figure 8A shows full blots, as example, with C1q detection by rabbit polyclonal anti-C1 q, anti-DNP mAb (indicated by “DNP”), and anti-C1q mAb 3C3. Figure 8B shows blots for all anti-C1q mAbs, and relevant parts were aligned as summary. To investigate the importance of conformational structure of C1q for binding by anti-C1q mAbs, Western blots loaded with C1 q in different levels of unfolding (heated and reduced, heated only and unheated) were stained with anti-C1q mAbs. Full blots shown as example demonstrate no background binding (Figure 8A). A strong staining of a band on the blot was observed on unheated C1 q for all anti-C1q mAbs (Figure 8B). On heated samples of C1 q, with or without reducing agent, none of the anti-C1q mAbs yielded any signal. In contrast, a polyclonal antibody detecting C1 q (indicated by “polyclonal” or “poly” in the Figures) showed clear and discrete bands at the expected molecular weights for all conditions. These results highlight the importance of the conformation of C1 q to allow anti-C1 q binding.

[0431] Figure 9A shows binding of anti-C1 q monoclonal antibodies to distinct non-competing epitopes on solid-phase C1 q. Each of the human anti-C1 q monoclonal antibodies designated 2D2, 1 F4, 4C11 , 4D2, 4E6, 4F5, 1 F5, 3C3, and 12F6, as well as a negative control mouse anti-C1 q monoclonal antibody 4A4B11 , were assessed for competition for binding to solid-phase C1q. Biotinylated antibodies of each monoclonal antibody were generated and were incubated together with nonbiotinylated antibodies as competitors in the presence of solid-phase C1q. Data are presented with the binding of only the biotinylated antibody set at 100%. The human anti-C1q antibodies are clearly different from the mouse anti-C1q negative control antibody as they do not display cross-inhibition. It can be further appreciated that due to the overall lack of cross-inhibition between the different human antibodies, distinct non-competing epitopes are bound by the different human antibodies.

[0432] Figure 9B shows anti-C1q mAbs binding to full C1q, C1q Collagen Like Region (CLR), and C1 q (recombinant) globular head (gh) domains. In order to map the binding of anti-C1 q mAbs to the different regions of the C1 q molecule, binding to CLR and recombinant globular head domain was evaluated. All anti-C1 q mAbs evidently bind to the CLR and not the globular head when tested in ELISA, with the exception of mAb 12F6, for which no targeted region could be identified.

[0433] Figure 10 shows uptake or binding of IgM- and C1q-labelled beads by THP-1 cells differentiated into macrophages. Figure 10A shows cell interaction with lgM / C1 q coated beads after incubation with anti-C1q mAbs. It can be seen that addition of 1 pg / ml anti-C1 q mAb could increase binding of the beads to the differentiated THP-1 cells as much as 5-fold. Figure 10B shows cell interaction with lgM / C1q coated beads after incubation with selected anti-C1q mAbs in the presence of a Fc blocking agent. The enhancement of uptake by anti-C1q mAb as shown in Figure 10A could be inhibited by pre-incubating the cells with an Fc receptor blocking agent. The combined data reveal that lgG1 anti-C1q mAbs might improve the ability to clear particles opsonized by IgM by phagocytosis.

[0434] Figure 11 shows electron tomography images of C1q bound by anti-C1q mAb. Anti-C1q binding was visualized on C1 / lgM complexes on liposomes with negative stain electron tomography, and on C1 q / lgG complexes with cryo-electron tomography. Anti-C1q antibody, bispecific antibody 1 F5xb12 (as also used in the solid-phase C1 q inhibition ELISA of Figure 6) was employed to be certain that the anti-C1q mAb could not bind C1 q through a second Fab arm or its Fc domain. It can be observed that the anti-C1q antibodies bind to C1q near the globular head region, and not on the top of the C1q molecule where all chains come together. It is further obvious that multiple anti-C1q antibodies bind to the same C1q protein, demonstrating the presence of multiple epitopes on one molecule. Figure 11 A shows 4.5 nm thick negative stain electron tomogram slices of C1q (blue) bound to hexameric anti-CD52 IgG (red) with anti-C1q bsAbs (orange) bound to C1 . Figure 11 B shows 8 nm thick cryo-electron tomogram slices of C1 (blue) bound to anti-CD52 IgM (green) with anti-C1q bispecific Abs (orange) bound to C1 on the surface of CD52 liposomes. First and third rows show tomogram slices; second and fourth rows show the same tomogram slices with complex map overlaid. Figure 12 shows binding of antibodies from SLE patient plasma, healthy anti-C1q negative serum, a mix of anti-C1q mAbs or polyclonal Rabbit anti-C1q (DAKO; A0136) to linear 21-mer C1q peptides. In an effort to further specify the location on C1q recognized by the anti-C1q mAbs, a set of 79 overlapping peptides was produced, with each peptide 21 amino acids long and 12 amino acids overlap between peptides, to cover all three chains of the C1q protein. Interestingly, no meaningful binding signal was observed for the anti-C1 q mAbs or anti-C1q positive plasma of 3 SLE patients on any of the peptides. As detection with a polyclonal anti-C1 q antibody did show reactivity for a number of peptides, the inability of the human anti-C1q autoantibodies to bind any of the peptides, together with the western blot data, points toward recognition of a structural epitope as opposed to an epitope consisting of a linear peptide.

[0435] Figure 13 shows that anti-C1 q mAbs increase Fc receptor engagement, but not complement activation, on immune complexes. Figure 13A shows complement activation on hexameric IgG complexes with or without extra C1q in ELISA detected on the level of C5b9. On the basis of the NHS titration, 1 % NHS was chosen for the experiment shown in Figure 13B to investigate the effect of anti- 01 q mAb. Figure 13B shows C5b9 deposition on hexameric IgG complexes in ELISA, in the presence of different anti-C1q mAb at 1 % NHS. Figure 13C shows binding of biotinylated FcRIIIa to hexameric IgG complexes with or without extra C1q in ELISA. On the basis of the FcRIIIa titration, 3 pg / ml FcRIIIa was used in the experiment shown in Figure 13D to investigate the effect of anti-C1 q mAb. Figure 13D shows that FcRIIIa-biotin binds to hexameric IgG complexes and different anti-C1q mAb in ELISA. Figure 13E shows that PMA-differentiated THP-1 cells bind lgM-coated / C1q opsonized beads, in the presence of anti-C1 q mAb. Figure 13F shows that human polymorphonuclear leukocytes phagocytose S. aureus bacteria opsonized with anti-WTA lgG4 and C1q, in the presence of anti-C1q mAb. For Figures 13B, D, E and F representative experiments from 3 repetitions are shown. For Figures 13B and D, each anti-C1q mAb was compared to anti-DNP with one-way ANOVA with Dunnett's multiple comparisons test. * P < 0.05; ** P < 0.01 ; *** P < 0.001 ; **** P < 0.0001 .

[0436] Figure 14 shows that the epitopes targeted by the anti-C1 q mAbs constitute the same epitopes targeted by anti-C1 q autoantibodies present in SLE sera. Figure 14 shows competition for binding to C1 q-coated ELISA, between biotinylated anti-C1q mAbs and polyclonal anti-C1 q antibodies from SLE serum, the percentage residual signal compared to no competition is indicated in the heatmap. Competition patterns were clearly split between the two groups identified in mAb-mAb competition. Epitopes targeted by SLE A and SLE B overlap with epitopes of mAbs 1 F4, 1 F5, 3C3 and 12F6, while SLE C antibodies compete with mAbs 2D2, 4C11 , 4D2, 4E6 and 4F5. Data is representative of two independent experiments. 100 indicates no competition, 0 indicates full competition.

[0437] General definitions

[0438] “Isolated” herein when referring to a polypeptide according to invention is understood to mean that the polypeptide is free from at least one component or all components as it is found in its natural environment. Such components include for example cells, proteins, nucleic acids, lipid membranes and nutrients. A polypeptide according to the invention can also be free or substantially free from all these components.

[0439] "Pharmaceutically acceptable" refers to a material that is not biologically or otherwise undesirable, i.e., the material may be administered to an individual along with the selected binding protein without causing any undesirable biological effects or interacting in a deleterious manner with any of the other components of the pharmaceutical composition in which it is contained.

[0440] Excipients are natural or synthetic substances formulated alongside an active ingredient (e.g., a polypeptide provided herein), included for the purpose of bulking-up the formulation or to confer a therapeutic enhancement on the active ingredient in the final dosage form, such as facilitating drug absorption or solubility. Excipients can also be useful in the manufacturing process, to aid in the handling of the active substance concerned such as by facilitating powder flowability or non-stick properties, in addition to aiding in vitro stability such as prevention of denaturation over the expected shelf life. Pharmaceutically acceptable excipients are well known in the art. A suitable excipient is therefore easily identifiable by one of ordinary skill in the art. By way of example, suitable pharmaceutically acceptable excipients include water, saline, aqueous dextrose, glycerol, ethanol, and the like.

[0441] Adjuvants are pharmacological and / or immunological agents that modify the effect of other agents in a formulation. Pharmaceutically acceptable adjuvants are well known in the art. A suitable adjuvant is therefore easily identifiable by one of ordinary skill in the art.

[0442] Diluents are diluting agents. Pharmaceutically acceptable diluents are well known in the art. A suitable diluent is therefore easily identifiable by one of ordinary skill in the art.

[0443] Carriers are non-toxic to recipients at the dosages and concentrations employed and are compatible with other ingredients of the formulation. The term “carrier’’ denotes an organic or inorganic ingredient, natural or synthetic, with which the active ingredient is combined to facilitate the application. Pharmaceutically acceptable carriers are well known in the art. A suitable carrier is therefore easily identifiable by one of ordinary skill in the art.

[0444] The term "variable region" refers to the region of an immunoglobulin that comprises one or more Ig domains substantially encoded by any of the VK, VA, and / or VH genes that make up the kappa, lambda, and heavy chain immunoglobulin genetic loci respectively. More specifically, the term refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to an antigen. The variable region of an immunoglobulin is therefore typically made up of two variable domains (i.e., the variable domain of the heavy chain and the variable domain of the light chain). The variable domains of the heavy chain and light chain (VH and VL, respectively) of a native antibody generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three hypervariable regions (HVRs). (See, e.g., Kindt et al. Kuby Immunology, 6th ed., W.H. Freeman and Co., page 91 (2007).). A single VH or VL domain may be sufficient to confer antigen-binding specificity. The term "hypervariable region" refers to the amino acid residues of an antibody which are responsible for antigen-binding. The hypervariable region generally comprises amino acid residues from a "complementarity determining region" or "CDR" (e.g. residues 24-34 (L1), 50-56 (L2) and SOO (L3) of the light chain variable domain and 31-35 (H1), 50-65 (H2) and 05-102 (H3) of the heavy chain variable domain according to Kabat et al, Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1001)) and / or those residues from a "hypervariable loop" (e.g. residues 26-32 (L1), 50-52 (L2) and 01-06 (L3) in the light chain variable domain and 26-32 (H1), 53-55 (H2) and 06-101 (H3) in the heavy chain variable domain according to Chothia and Lesk J. Mol. Biol. 106:001-017 (1087)).

[0445] The amino-terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition, generally referred to in the art and herein as the "Fv domain" or "Fv region". In the variable region, three loops are gathered for each of the V domains of the heavy chain and light chain to form an antigen-binding site. Each of the loops is referred to as a complementarity-determining region (hereinafter referred to as a "CDR"), in which the variation in the amino acid sequence is most significant. "Variable" refers to the fact that certain segments of the variable region differ extensively in sequence among antibodies. Variability within the variable region is not evenly distributed. Instead, the V regions consist of relatively invariant stretches called framework regions (FRs) of 15-30 amino acids separated by shorter regions of extreme variability called "hypervariable regions" that are each 9-15 amino acids long or longer. Each VH and VL is composed of three hypervariable regions ("complementary determining regions," "CDRs") and four FRs, arranged from amino- terminus to carboxy-terminus in the following order: FR1 -CDR1 - FR2-CDR2-FR3- CDR3-FR4.

[0446] An antibody may comprise at least two variable regions that specifically bind to human solidphase C1q. Each variable region may comprise the CDRs of a light chain variable domain (VL) (i.e., VL CDR1 , VL CDR2, and VL CDR3) and / or the CDRs of a heavy chain variable domain (VH) (i.e., VH CDR1 , VH CDR2, and VH CDR3). As stated elsewhere herein, the CDRs from one of VL or VH may be sufficient to confer antigen binding specificity. In other cases, antigen binding specificity may be obtained by the presence of CDRs 1 , 2, and 3, from both VL and VH.

[0447] Specific examples of combinations of CDRs that confer antigen binding specificity to human solid-phase C1q are provided below. The CDR sequences have been identified using IMGT / V-QUEST program version: 3.5.20 (14 September 2020) - IMGT / V-QUEST reference directory release: 202038- 1 (14 September 2020) (http: / / imgt.org / IMGT_vquest / vquest), selecting for homo sapiens sequences These CDR combinations form a variable region of an antibody (including a Fab2, Fab3, di-scFv, scFV-Fc, duobody, camelid antibody or minibody) that specifically binds to human solid-phase C1 q.

[0448] The term “antibody” or “antibodies” include monoclonal, polyclonal, chimeric, single chain, bispecific, human, and humanized antibodies as well as active multivalent fragments thereof. Examples of active multivalent fragments of molecules that bind to known antigens and are useful in the present invention include F(ab')2, F(ab')3, diabodies, triabodies, scFv-Fc and di-scFv and minibodies, including the products of a Fab immunoglobulin expression library and epitope-binding multivalent fragments of any of the antibodies and multivalent fragments mentioned above.

[0449] In a particular example, the antibody may be a monoclonal antibody. As used herein, the term “monoclonal antibody” refers to an antibody that is mass produced in the laboratory from a single clone and that recognizes only one antigen. Monoclonal antibodies may be generated by any appropriate technique known in the art (e.g., by production in HEK or insect cells, or by generation of B cell hybridomas).

[0450] As used herein, the term “chimeric antibody” refers to a monoclonal antibody comprising a variable region, i.e., binding region, from one source or species, (i.e., non-human primates, humans, dogs, cats, horses, cows, pigs, guinea pigs, mice, rats, and the like ) and at least a portion of a constant region derived from a different source or species, usually prepared by recombinant DNA techniques. Chimeric antibodies comprising a mouse variable region and a human constant region are examples. Such chimeric antibodies are the product of expressed immunoglobulin genes comprising DNA segments encoding mouse immunoglobulin variable regions and DNA segments encoding human immunoglobulin constant regions. Other forms of “chimeric antibodies” encompassed by the present disclosure are those in which the class or subclass has been modified or changed from that of the original antibody, for example to alter them so that they are complement and Fc receptor binding deficient. Methods for producing chimeric antibodies involve conventional recombinant DNA and gene transfection techniques now well known in the art. See, e.g., Morrison, S. L., et al., Proc. Natl. Acad Sci. USA 81 (1984) 6851-6855; U.S. Pat. No. 5,202,238 and U.S. Pat. No. 5,204,244.

[0451] In a particular example, the antibody may be a human antibody or a humanized antibody.

[0452] As used herein the term “humanized antibody” or “humanized version of an antibody” refers to antibodies in which the framework or “complementarity determining regions” (CDR) have been modified to comprise the CDR of an immunoglobulin of different specificity as compared to that of the parent immunoglobulin. In some examples, the CDRs of the VH and VL are grafted into the framework region of human antibody to prepare the “humanized antibody.” See e.g., Riechmann, L., et al., Nature 332 (1988) 323-327; and Neuberger, M. S., et al., Nature 314 (1985) 268-270. The heavy and light chain variable framework regions can be derived from the same or different human antibody sequences. Both the heavy chain and the light chain may be required for effective antigen binding. The human antibody sequences can be the sequences of naturally occurring human antibodies. Human heavy and light chain variable framework regions are listed e.g., in Lefranc, M.-P., Current Protocols in Immunology (2000) — Appendix 1 P A.1 P.1-A.1 P.37 and are accessible via IMGT, the international ImMunoGeneTics information System® (http: / / imgt.cines.fr) or via http: / / vbase.mrc- cpe.cam.ac.uk, for example. Optionally the framework region can be modified by further mutations. Exemplary CDRs correspond to those representing sequences recognizing the antigens noted above for chimeric antibodies. In some examples, such humanized version is chimerized with a human constant region. As used herein the term “human antibody” is intended to include antibodies having variable and constant regions derived from human germ line immunoglobulin sequences. Human antibodies are well-known in the state of the art (van Dijk, M. A., and van de Winkel, J. G., Curr. Opin. Chem. Biol. 5 (2001) 368-374). Human antibodies can also be produced in transgenic animals (e.g., mice) that are capable, upon immunization, of producing a full repertoire or a selection of human antibodies in the absence of endogenous immunoglobulin production. Transfer of the human germ-line immunoglobulin gene array in such germ-line mutant mice results in the production of human antibodies upon antigen challenge (see, e.g., Jakobovits, A., et al., Proc. Natl. Acad. Sci. USA 90 (1993) 2551-2555; Jakobovits, A., et al., Nature 362 (1993) 255-258; Brueggemann, M. D., et al., Year Immunol. 7 (1993) 33-40). Human antibodies can also be produced in phage display libraries (Hoogenboom, H. R., and Winter, G., J. Mol. Biol. 227 (1992) 381-388; Marks, J. D., et al., J. Mol. Biol. 222 (1991) 581-597). The techniques of Cole, A., et al. and Boerner, P., et al. are also available for the preparation of human monoclonal antibodies (Cole, A., et al., Monoclonal Antibodies and Cancer Therapy, Liss, A. R. (1985) p. 77; and Boerner, P., et al., J. Immunol. 147 (1991) 86-95).

[0453] In a particular example, the antibody may be selected from a Fab2, Fab3, di-scFv, scFv-Fc, duobody, or a minibody.

[0454] Other examples include single domain antibodies such as those found in camelids including, but not limited to, llamas and alpacas; and cartilaginous fish including, but not limited to, sharks which are widely known in the art.

[0455] As used herein “single chain antibody” refers to single chain Fv molecules (scFv), wherein a VH domain and a VL domain are linked by a peptide linker which allows the two domains to associate to form an antigen binding site (Bird et al., 1988, Science 242:423-426, Huston et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:5879-5883 or a bi-specific single chain Fv (WO 03 / 11161). Typical scFv linkers are well known in the art, are generally 10 to 25 amino acids in length and include glycines and serines.

[0456] Single domain antibody fragments (VHH or nanobodies) e.g., the functional antibodies produced by camelids that are devoid of light chains and wherein a single N-terminal domain is fully capable of antigen binding are also an example of a binding agent described herein. Such fragments can also form bivalent VHHH, or pentabodies (i.e. , with 5 VHH domains).

[0457] As used herein, a “di-ScFv” refers to a dimerized scFV.

[0458] As used herein, “minibodies” are minimized antibody-like proteins comprising a scFv joined to a CH3 domain. See Hu et al., 1996, Cancer Res. 56:3055-3061. In some cases, the scFv can be joined to the Fc region, and may include some or the entire hinge region.

[0459] By "Fab" or "Fab region" as used herein is meant the polypeptides that comprise the VH, CH1 , VL, and CL immunoglobulin domains. Fab may refer to this region in isolation, or this region in the context of a full-length antibody or antibody fragment or fab fusion protein.

[0460] The terms “Fab”, “Fab region”, “Fab portion” or “Fab fragment” are understood to define a polypeptide that includes a VH, a CH1 , a VL, and a CL immunoglobulin domain. Fab may refer to this region in isolation, or this region in the context of an antibody molecule according to the invention, as well as a full-length immunoglobulin or immunoglobulin fragment. Typically, a Fab region contains an entire light chain of an antibody. A Fab region can be taken to define “an arm” of an immunoglobulin molecule. It contains the epitope-binding portion of that Ig. The Fab region of a naturally occurring immunoglobulin can be obtained as a proteolytic fragment by a papain-digestion. A “F(ab')2 portion” is the proteolytic fragment of a pepsin-digested immunoglobulin. A “Fab' portion” is the product resulting from reducing the disulfide bonds of an F(ab')2 portion. As used herein the terms “Fab”, “Fab region”, “Fab portion” or “Fab fragment” may further include a hinge region that defines the C-terminal end of the antibody arm (cf. above). This hinge region corresponds to the hinge region found C- terminally of the CH1 domain within a full-length immunoglobulin at which the arms of the antibody molecule can be taken to define a Y. The term hinge region is used in the art because an immunoglobulin has some flexibility at this region.

[0461] By "Fc fusion" as used herein is meant a protein wherein one or more polypeptides is operably linked to Fc. Fc fusion is herein meant to be synonymous with the terms "immunoadhesin", "Ig fusion", "Ig chimera", and "receptor globulin" (sometimes with dashes) as used in the prior art (Chamow et al., 1996, Trends Biotechnol 14:52-60; Ashkenazi et al., 1997, Curr Opin Immunol 9:195-200). An Fc fusion combines the Fc region of an immunoglobulin with a fusion partner, which in general may be any protein, polypeptide, or small molecule. The role of the non-Fc part of an Fc fusion, i.e., the fusion partner, is to mediate target binding, and thus it is functionally analogous to the variable regions of an antibody. Virtually any protein or small molecule may be linked to Fc to generate an Fc fusion. Protein fusion partners may include, but are not limited to, the target-binding region of a receptor, an adhesion molecule, a ligand, an enzyme, a cytokine, a chemokine, or some other protein or protein domain. Small molecule fusion partners may include any therapeutic agent that directs the Fc fusion to a therapeutic target. Such targets may be any molecule, e.g., an extracellular receptor that is implicated in disease.

[0462] “Immunotherapy” as used herein refers to the treatment of disease by activating, enhancing, suppressing or inhibiting the immune system.

[0463] As used herein the term “antibody fragments” refers to a portion of a full-length antibody, for example possibly a variable domain thereof, or at least an antigen binding site thereof. Examples of antibody fragments include diabodies, single-chain antibody molecules, and multispecific antibodies formed from antibody fragments. scFv antibodies are, e.g., described in Huston, J. S., Methods in Enzymol. 203 (1991) 46-88. Antibody fragments can be derived from an antibody of the present invention by a number of art-known techniques. For example, purified monoclonal antibodies can be cleaved with an enzyme, such as pepsin, and subjected to HPLC gel filtration. The appropriate fraction containing Fab fragments can then be collected and concentrated by membrane filtration and the like. For further description of general techniques for the isolation of active fragments of antibodies, see for example, Khaw, B. A. et al. J. Nucl. Med. 23:1011-1019 (1982); Rousseaux et al. Methods Enzymology, 121 :663-69, Academic Press, 1986. Antibody fragments useful to the invention are multivalent antibody fragments. As used herein, the term “trigger” is understood to be synonymous with “induce”.

[0464] Throughout the description and claims of this specification, the words “comprise” and “contain” and variations of them mean “including but not limited to”, and they are not intended to (and do not) exclude other moieties, additives, components, integers, or steps.

[0465] Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0466] Features, integers, characteristics, compounds, chemical moieties, or groups described in conjunction with a particular aspect, embodiment or example of the invention are to be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith.

[0467] The patent, scientific and technical literature referred to herein establish knowledge that was available to those skilled in the art at the time of filing. The entire disclosures of the issued patents, published and pending patent applications, and other publications that are cited herein are hereby incorporated by reference to the same extent as if each was specifically and individually indicated to be incorporated by reference. In the case of any inconsistencies, the present disclosure will prevail.

[0468] Unless defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. For example, Singleton and Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d Ed., John Wiley and Sons, NY (1994); and Hale and Marham, The Harper Collins Dictionary of Biology, Harper Perennial, NY (1991) provide those of skill in the art with a general dictionary of many of the terms used in the invention. Although any methods and materials similar or equivalent to those described herein find use in the practice of the present invention, the preferred methods and materials are described herein. Accordingly, the terms defined immediately below are more fully described by reference to the Specification as a whole. Also, as used herein, the singular terms "a", "an," and "the" include the plural reference unless the context clearly indicates otherwise. Unless otherwise indicated, nucleic acids are written left to right in 5' to 3' orientation; amino acid sequences are written left to right in amino to carboxy orientation, respectively. It is to be understood that this invention is not limited to the particular methodology, protocols, and reagents described, as these may vary, depending upon the context they are used by those of skill in the art.

[0469] All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and / or steps are mutually exclusive.

[0470] Each feature disclosed in this specification (including any accompanying claims, abstract and drawings), may be replaced by alternative features serving the same, equivalent, or similar purpose, unless expressly stated otherwise. Thus, unless expressly stated otherwise, each feature disclosed is one example only of a generic series of equivalent or similar features.

[0471] The invention is not restricted to the details of any foregoing embodiments. The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.

[0472] Aspects of the invention are demonstrated by the following non-limiting examples.

[0473] Examples

[0474] Example 1

[0475] Materials and methods

[0476] Isolation ofanti-C1q B cells by FACS

[0477] Peripheral blood and serum was collected from anti-C1q-positive donors after obtaining their informed consent (B19.008 / AB / ab). Peripheral blood mononuclear cells (PBMC) were isolated using Ficoll- Paque (LUMC pharmacy) gradient centrifugation and B cells were enriched using the EasySep Human B cell isolation kit (Stem Cell Technologies) following instructions provided by the manufacturer. To optimize the anti-C1q staining procedure, HEK cells were transduced with lentiviral vectors encoding a transmembrane domain with heavy and light chains of mouse anti-C1q clone JL-1 , under selection of GFP production and puromycin resistance respectively, as described before (Kerkman, Ann Rheum Dis, 2016; Trouw, J Clin Invest, 2004). The resulting cells mimic B cells with a C1q-reactive B-cell receptor. Staining complexes for solid-phase C1 q consisted of purified C1q (Complement Technologies), pre-formed hexamers of anti-DNP lgG1-RGY antibodies, DNP-biotin peptides (LUMC peptide facility) and streptavidin-PE (Bio Rad) or streptavidin-AlexaFluor647 (Invitrogen) (Figure 1A). lgG1 antibodies (clone G2a-2) directed against the hapten dinitrophenol (DNP) (Gonzalez, Mol Immunol, 2003) were engineered with hexamerization-enhancing mutations E345R, E430G, and S440Y to bring C1 q in its solid-phase conformation and produced in-house (Ugurlar, Science, 2018; Wang, mol cell, 2016). A small peptide consisting of DNP-glycine, a PEG2-GSGSGSG linker and biotin served as a bridge between the hexamer of anti-DNP lgG1 and streptavidin-coupled fluorochromes.

[0478] B cells were incubated with solid-phase C1 q staining complexes for 45 minutes at 4°C. After washing in PBS / 1% fetal calf serum (FCS), B cells were further stained with mouse anti-human CD27-FITC (ThermoFisher; clone CLB-27 / 1), CD3-Pacific Blue (BD Biosciences; clone SP34-2) and lgD-PE-Cy7 (BD Biosciences, clone IA6-2) and incubated for 45 minutes at 4°C. Single B cells detected as CD3_, IgD-, CD27+and double positive for C1q-complex staining were sorted on a FACSAria III Cell Sorter (BD Biosciences) and collected at one cell per well in a 96-well flat bottom plates (Corning). The wells contained 10A5 irradiated EL4B5 cells expressing CD40L in 200 pl IMDM (Lonza) supplemented with 10% FCS, 2mM L-glutamine (Gibco), 50 pM 2-mercaptoethanol (Sigma-Aldrich), 100 U / ml penicillin, 100 pg / ml streptomycin (both Gibco), 20 pg / ml Insulin-transferrin-sodium selenite (Sigma-Aldrich), 50 ng / ml IL-21 (Gibco), 1 ng / ml IL-1 p (Miltenyi Biotec), 0.3 ng / ml TNFalpha (Miltenyi Biotec) and 0.5 pg / ml R848 (Sigma-Aldrich). Plates with B cells were incubated for 13 days at 37°C and 5% CO2 before analysis.

[0479] Identification and genetic analysis of anti-C1q producing B cells

[0480] Supernatant of sorted B cells was harvested after a 13-day expansion period and screened for IgG production and C1 q specificity by in-house enzyme-linked immunosorbent assay (ELISA). B-cell clones positive for anti-C1q production were lysed and RNA was isolated using TriZol reagent (Invitrogen). Subsequently, cDNA was synthesized using PrimeScript Reverse Transcriptase (Takara) followed by RACE PCR to amplify variable domains of the heavy (VH) and kappa or lambda light (VL) chain. Isolated VH and VL fragments were ligase-independently cloned into pcDNA3.3 plasmids containing lgG1 , kappa or lambda constant domains. The plasmids were then sequenced by the Leiden Genome Technology Center (LGTC) to obtain the VH and VL sequences. Sequences were analysed for VDJ gene usage and complementarity-determining region (CDR) identification by IMGT V-quest (V-quest: Brochet et al., Nucl Acids Res, 2008).

[0481] Production and purification of monoclonal antibodies

[0482] Heavy and light chain plasmids were co-transfected in Expi293F cells using ExpiFectamine, Opti- MEM, and Expi293 expression medium (all ThermoFisher) according to the manufacturer’s instructions. Supernatant was harvested after 5-7 days, filtered and antibodies were purified on protein A resin (Genscript). Subsequently, buffer was exchanged to PBS and antibodies were concentrated on 50 kDa Amicon centrifugal filters (Merck Millipore). Antibody concentration was measured using an in-house sandwich ELISA as described previously (Heidt, Clin Exp Imm, 2010).

[0483] ELISA to screen individuals foranti-C1q positivity

[0484] Serum of selected individuals was screened for the presence of anti-C1q antibodies by QUANTA Lite Anti-C1q ELISA (Inova Diagnostics) according to the manufacturer’s protocol. Briefly, samples were diluted 1 :101 in Sample Diluent and 100 pl diluted samples or supplied controls was added to the wells. After 30-minute incubation, wells were washed and incubated with 100 pl HRP IgG conjugate for another 30 minutes. Wells were washed, stained for 30 minutes with 100 pl TMB Chromogen while protected from light, then 100 pl HRP Stop Solution was added. Absorbance at 450 nm was measured and used to calculate anti-C1 q units based on positive controls samples provided in the kit. The cutoff for positivity was 20 units, as recommended by the manufacturer.

[0485] ELISA for binding ofanti-C1q mAbs to C1q on natural ligands and polyreactivity

[0486] ELISA was performed to evaluate reactivity of the isolated mAbs towards C1q on natural ligands and towards common targets of polyreactive antibodies. Nunc MaxiSorp plates (ThermoFisher) were coated with 10 pg / ml calf thymus single stranded DNA (Sigma-Aldrich), LPS (Sigma-Aldrich), pre-pro- insulin (produced in-house), intravenous immunoglobulins (IVIG; Sanquin), CRP (Calbiochem), 5 pg / ml IgM (Sigma-Aldrich) or 5 pg / ml C1q in bicarbonate coating buffer (0.1 M Na2COs / NaHCO3, at pH 9.6) for 1 hour at 37°C. Plates were washed 3 times with PBS / 0.05% Tween after every incubation. Plates were blocked with 100 pl / well PBS / 1 % BSA for 1 hour at 37°C. Wells coated with IVIG, IgM or CRP were incubated with 5 pg / ml C1 q diluted in PBS / 0.05% Tween / 1 % BSA (hereafter abbreviated to PTB) for 1 hour at 37°C. After washing, plates were incubated with anti-C1 q mAbs biotinylated with Pierce Antibody Biotinylation Kit (ThermoFisher) diluted in PTB and incubated for 1 hour at 37°C. Binding of anti-C1q mAbs was detected by horseradish peroxidase (HRP)-coupled streptavidin (ThermoFisher) in PTB, incubated for 1 hour at 37°C. After the final washing sequence, 50 pl 2,2'- azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) / 0.015% H2O2 (both from Merck) was added and absorbance at 415 nm was measured using a microplate reader (Bio-Rad iMark).

[0487] ELISA for fluid phase C1q inhibition to anti-C1q

[0488] ELISA plates were coated with 10 pg / ml C1q in coating buffer overnight at 4°C and subsequently blocked with PBS / 1%BSA. Biotinylated anti-C1q mAbs were mixed with 20 pg / ml C1q, 20 pg / ml C1 (Complement Technologies), 5% or 25% pooled normal human serum (NHS) or 5% or 25% C1q depleted serum (Complement Technologies), in PTB containing 0.5 M NaCI to prevent C1 q-Fc interactions (Kohro-Kawata, J Rheumatol, 2002). Samples with serum were also supplemented with 10 mM EDTA to prevent complement activation. After pre-incubation on ice for 30 minutes, samples were incubated in the C1 q-coated wells for 1 hour at 37°C. Anti-C1q mAbs were detected with streptavidin-HRP, plates were then developed by incubating with ABTS / 0.015% H2O2 and measured using a microplate reader. Percentage signal was calculated by setting the absorbance values of samples without C1q competition at 100% for each individual mAb. Antibody concentrations for summary figures were chosen to be near the saturation point for individual mAbs, being 160 ng / ml (1 F5, 4F5, 4A4B11 ), 640 ng / ml (3C3, 4D2, 4E6), 2560 ng / ml (1 F4, 2D2, 4C11 ) or 10240 ng / ml (12F6).

[0489] ELISA to determine inhibition ofanti-C1q binding to solid-phase C1q

[0490] For clones 1 F4, 1 F5 and 4F5, antibodies were produced with containing LALAPG mutations to render their Fc domain functionally inactive (LALA-PG [Lo, J Biol Chem, 2017]) and to allow for Fab-arm exchange. These antibodies were then combined with anti-HIV gp120 clone b12 through Fab-arm exchange following the protocol described by Labrijn et al (Labrijn, Nature protocols, 2014). The resulting bispecific antibodies (bsAbs) were functionally monovalent for C1 q binding, with inactive Fc domains to specifically investigate the interactions between one C1q molecule and one antigen binding domain. ELISA plates were coated with 10 pg / ml C1 q in coating buffer overnight at 4°C and subsequently blocked with PBS / 1%BSA. Biotinylated anti-C1q bsAbs in PTB were mixed with C1q or human serum albumin (HSA) coupled to Carboxyl Fluorescent Particles, 0.4-0.6 pm (Spherotech), or C1 q or HSA in solution. After pre-incubating of 30 minutes, each well in the ELISA plate received 0.1 g anti-C1 q antibody and 1 pg C1q or HSA, which in case of solid-phase competition was coupled to 109beads. After incubating for 1 hour at 37°C, plates were washed and bound, biotin-labelled antiCi q antibodies were detected with streptavidin-HRP. Plates were developed by incubating with ABTS / 0.015% H2O2 and measured using a microplate reader.

[0491] Flow cytometry of opsonized and necrotic cells

[0492] PBMC’s were isolated from healthy donor peripheral blood by Ficoll-Paque gradient centrifugation. Cells were either opsonized by antibodies by incubating with recombinant anti-CD52 (Alemtuzumab) lgG1 containing hexamer-enhancing mutations RGY (Diebolder, Science, 2014) for45 minutes at 4°C or made necrotic by incubating for 30 minutes at 56°C. Cells were then seeded into 96-wells plates at 100 000 cells per well, unbound anti-CD52 antibody was washed away two times by adding 200 pl FACS buffer (PBS / 2% FCS), centrifuging and removing the supernatant. C1 q was added at 2 pg / ml in FACS buffer and incubated with the cells for 45 minutes at 4°C. After washing, cells were incubated with 10 pg / ml biotinylated anti-C1 q mAbs for 45 minutes at 4°C and washed again. Binding of antiCi q mAbs to the cells was detected by incubating with 2 pg / ml streptavidin-AlexaFluor647 for 45 minutes at 4°C. As positive control for C1 q binding, polyclonal rabbit anti-C1q (DAKO) and BrilliantViolet421 -labelled donkey anti-rabbit IgG (BioLegend) were used. After the final wash, fluorescent signal on the cells was measured on a FACSCanto flow cytometer (BD Biosciences).

[0493] Western blots of C1q in various conformational states

[0494] C1 q in different conformational states was prepared in Laemmli sample buffer (Bio-Rad) as either unheated, heated for 5 minutes at 95°C, or heated and reduced by adding 5% 2-mercaptoethanol in the sample buffer. A total of 2 pg C1q was applied per lane of a 4-15% gradient precast polyacrylamide gel (Bio-Rad) and run at 110 V for 1 hour. The PageRuler Plus Prestained Protein Ladder, 10 to 250 kDa (ThermoFisher) served as reference for the molecular weights of the bands.

[0495] The gel was washed in ultrapure water and protein was blotted to a nitrocellulose membrane using a T rans-Blot Turbo T ransfer System (both Bio-Rad). The blots were washed 3 times with ultrapure water and blocked with 30 mg / ml skim milk powder (Fluka) in PBS / 0.05% Tween for 90 minutes. After washing 2 times with ultrapure water, the blots were cut into 3 pieces to be stained with different antibodies, and incubated with 5 pg / ml anti-C1 q mAb or 1 :20000 rabbit anti-C1q polyclonal antibody in blocking buffer for 1 hour. The blots were washed 3 times with PBS / 0.05% Tween, then incubated with rabbit anti-human IgG-HRP (DAKO), or with 1 :20000 goat anti-rabbit-HRP (DAKO) matching the primary antibodies. After 1 hour, blots were washed 3 times with PBS / 0.05% Tween and once with ultrapure water. Blots were then stained with ECL blotting reagent (Cytiva) and imaged on ChemiDoc MP Imaging System (Bio-Rad).

[0496] ELISA to detect binding ofanti-C1q to C1q collagen-like region and globular heads ELISA plates were coated with C1 q CLR made by limited proteolysis of purified serum C1q, recombinant single chain globular head domains [described in Moreau, Front Immunol, 2016] (both gifts from Nicole Thielens), or purified intact C1 q at 10 pg / ml in coating buffer for 1 hour at 37°C. The plates were then blocked, washed and incubated with 4 pg / ml anti-C1 q mAb, or 10 pg / ml for mAb 12F6, for 1 hour at 37°C. After washing, bound anti-C1 q mAb was detected with rabbit anti-human IgG-HRP and plates were developed by incubating with ABTS / 0.015% H2O2 and measured using a microplate reader. Absorbance values for binding to intact C1 q were set to 100% for each mAb to facilitate easier comparison.

[0497] ELISA for competition between anti-C1q mAbs and purified SLE antibodies

[0498] ELISA plates were coated with 10 pg / ml C1 q in coating buffer overnight at 4 °C and subsequently blocked with PBS / 1%BSA for 1 hour at 37 °C. Competitor anti-C1q mAb at 64 pg / ml final concentration, or protein A-purified SLE antibodies at 5 mg / ml, was added to the plates in PBS / 0.05% Tween / 1% BSA (PTB) and incubated 1 hour at 37 °C. Without washing, biotinylated anti-C1q mAb was then added at concentration between 0.15-12 pg / ml depending on the concentration needed to obtain near saturation binding signal without competition. After incubation for 1 hour at 37 °C, plates were washed and biotinylated anti-C1q mAb binding was detected by 0.1 pg / ml streptavidin-HRP. Plates were developed by incubating with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid / 0.015% H2O2 and absorbance was measured at 415 nm using a microplate reader. Percentage residual binding was calculated by dividing absorbance in presence of competition by absorbance in absence of competition.

[0499] ELISA for binding ofanti-C1q to C1q peptides

[0500] A comprehensive set of peptides covering the A, B and C chains of C1 q was produced at the in-house peptide facility of the LUMC. Peptides were 21 amino acids long, overlapped 12 amino acids with the next peptide, and were C-terminally biotinylated after a aminohexanoic acid linker. Additionally, all cysteine residues were converted to a-aminobutyric acid (Abu), which better resembles a cysteine in disulphide bond as it would be in the full C1q protein. Peptides were initially dissolved in DMSO, then diluted 1000x in PTB to 10 pg / ml and incubated on streptavidin coated plates (ThermoFisher) for 1 hour at 37°C. After washing, plates were incubated with a mix of the anti-C1 q mAbs (5 pg / ml for each mAb), a mix of anti-C1q positive plasma samples (each 1 :50 diluted), a mix of 3 samples from antiCi q negative individuals or 1 :1000 rabbit anti-C1q in PTB buffer for 1 hour at 37°C. After washing, bound antibodies were detected with matched rabbit anti-human IgG-HRP or goat anti-rabbit-HRP for the polyclonal and plates were developed by incubating with ABTS / 0.015% H2O2 and measured using a microplate reader.

[0501] Flow cytometry of differentiated THP-1 cells binding opsonized beads The THP-1 cell line (ATCC TIB-202) was cultured in RPMI (Gibco) with 10% FCS, 2mM L-glutamine, 100 U / ml penicillin and 100 pg / ml streptomycin (all Gibco). Cells were differentiated to a macrophage phenotype by incubating 100000 cells / well in a 48 wells plate with phorbol 12-myristate 13-acetate (Sigma-Aldrich) in the medium. After 3 days incubating at 37°C, medium was replaced by PMA-free medium and cells were incubated for a further 5 days. Yellow fluorescent 0.46 pm carboxyl beads (Spherotech) were loaded with I g M , washed 3 times in PBS, and incubated with C1 q for 1 hour at 4°C. After washing, beads suspension was incubated with anti-C1 q mAb in RPMI for 1 hour at 4°C, after which the mixture was added to the differentiated THP-1 cells. Per well, 3 x 10A8 beads, incubated with 0.1 pg IgM, 0.5 pg C1 q and 0.2 pg anti-C1q mAb were added. For experiments with Fc blocking reagent, cells were incubated with 10 pg / ml Human Fc block (BD Biosciences) for 10 minutes at room temperature immediately before adding the beads suspension. Plates were centrifuged for 1 minute at 100 g to bring the beads in contact with the cells and were then incubated for 1 hour at 37°C to allow binding and phagocytosis of anti-C1q mAb-covered beads by the cells. The beads suspension was removed and the cells were treated with trypsin (Gibco) to detach them from the plate. Cells were resuspended, washed in FACS buffer and then analyzed on a FACSCanto flow cytometer. Binding or phagocytosis was measured by FITC fluorescence. Differentiation of THP-1 cells was confirmed by increased expression of CD11 b (by mouse anti-CD11 b-APC, clone D12, BD Biosciences) and decreased expression of CD15 (by mouse anti-CD15-BV510, clone W6D3, BD Biosciences).

[0502] Avidity measurement ofanti-C1q mAbs by surface plasmon resonance

[0503] Surface plasmon resonance (SPR) was employed to determine the affinity of anti-C1q mAbs for Cl q, using a Biacore T200 machine (Cytiva). Biotinylated C1 q was immobilized on a streptavidin coated chip (Cytiva) to a response of 146.4 response units, with an empty channel for compensation. Next, titration curves of bivalent anti-C1q mAb in PBS / 0.05% Tween / 0.5 mg / ml BSA from 40 to 0.05 pg / ml were prepared. These samples were stored in the Biacore machine at 4°C and covered with a breakable seal during the measurement to ensure a stable quality of the samples. Samples were run in increasing concentration steps, with chip regeneration by 2 times 30 seconds flowing 10 mM glycine, pH=2.0. Resulting binding curves were analysed using Biacore T200 Evaluation Software 3.2.1 (Cytiva) to fit a 1 :1 binding model, resulting in association and dissociation constants for each antiCi q mAb.

[0504] Sample preparation and data collection for negative stain electron tomography

[0505] C1 q (0.270 mg / ml final concentration) was incubated with lgG1-anti-CD52-RGY (0.540 mg / ml final concentration) for 30 min at 4 °C. Next, anti-C1q bsAb 1 F5xb12 (made as described for the solidphase C1q inhibition ELISA; used at 0.090 mg / ml final concentration) was added and samples were incubated for another 30 min at 4 °C. Samples were purified using a Superdex 200 Increase 3.2 / 300 column (Cytiva). Column was equilibrated with PBS on an Akta pure system (Cytiva). Size exclusion fractions were diluted 1 :10 in water and loaded on freshly plasma-cleaned 200 mesh carbon coated copper grids (Electron Microscopy Sciences) and incubated for 1 min, before blotting using Whatman paper. Samples were stained using 2% uranyl formate for 1 min.

[0506] Negative stain tilt-series were collected on a FEI Tecnai T12 Biotwin with LaB6 source, operating at 120 kV on a FEI Eagle 4k x 4k CCD camera. Tilt series were collected using Xplore 3D (Thermo Fisher Scientific) at 49,000x magnification and a pixel size of 4.546 A using a continuous acquisition scheme from -60° to +60° with a tilt increment of 3°. A total dose of 100 e7A2and a defocus of -4 pm was used. Tracking and focusing were performed before every third image acquisition.

[0507] Sample preparation and data collection for cryo-electron tomography

[0508] Liposomes with 1% mCD52 (CD52 mimotope peptide; amino acid sequence TSSPSAD, synthesized by Aimee Boyle, Leiden Institute of Chemistry, the Netherlands), composed of dimyristoylphosphatidylcholine (DMPC), dimyristoylphosphatidylglycerol (DMPG), cholesterol and mCD52-cholesterol (44:5:50:1 mol%) in PBS were produced as previously described (Sharp, PNAS, 2019; Lubbers, Clin Exp Imm, 2020; Sharp, Cell Rep, 2016). All lipids, except mCD52-cholestrol, were purchased from Avanti Polar Lipids.

[0509] Liposomes were incubated with anti-CD52 IgM (YTH66.9HL, Biorad; 0.075 mg / ml final concentration) at room temperature for 20 min. To be able to image C1 binding to an antibody platform, samples were cooled down to 4 °C for 20 min before normal human serum (NHS; 10% v / v final concentration) was added and then incubated for another 20 min at 4 °C. Afterwards, anti-C1 q bsAb 1 F5xb12 (0.075 mg / ml final concentration) was added and incubated for 20 min at 4 °C. As fiducial markers, pre-cooled 5 nm gold colloids coated with bovine serum albumin were added. Samples were applied to freshly plasma cleaned 200 mesh lacey carbon support copper grids (Electron Microscopy Sciences). Grids were loaded into a Leica EMGP (Leica Microsystems) and incubated for 60 sec at 4 °C and 80% humidity, followed by blotting for 1.5 seconds from the back and vitrification in liquid ethane.

[0510] A Talos Arctica (Thermo Fisher Scientific) operated at 200 kV with a K3 direct electron detector, in counted-mode and a Bioquantum energy filter (Gatan) with a slit width of 20 eV was used for data collection. Tilt series were collected using Tomography 5.5.0 (Thermo Fisher Scientific) at 49,000x magnification, equal to a pixel size of 1 .74 A using a dose symmetric scheme from 0 to + / - 57°, in 3° increments. A total dose of 60 e7A2was used. Tilt series were collected with a defocus between -4 and -6 pm. Tracking and focusing were performed before each image acquisition.

[0511] Tomogram reconstruction

[0512] Alignment of cryo-electron tomography raw frames was performed using the “alignframes” command from the software program IMOD 4.11 .13 (Kremer, J Struct Biol, 1996). Additionally, IMOD was used to reconstruct negative stain, as well as cryo-electron tomograms using weighed back projected with a SIRT-like filter equivalent to 5 iterations. Negative stain tomograms were aligned using patch tracking, whereas cryo-electron tomograms were aligned using a seed model. Cryo-electron tomograms were binned by 8 before further image analyses. lgM / C1 / 1 F5xb12 maps, as well as lgG1 - CD52-RGY / C1q / 1 F5xb12 maps were displayed on the tomographic slices using UCSF Chimera 1.16 (Pettersen, J Comput Chem, 2004).

[0513] Results

[0514] Isolation and verification of monoclonal anti-C1q from peripheral blood B cells

[0515] To select possible donors for anti-C1q reactive B cells, serum of SLE patients and healthy donors was screened for the presence of anti-C1q antibodies. For healthy donors, 9 of 16 selected serum samples were positive for anti-C1 q, while 11 out of 14 serum samples from SLE patients were positive. Single B cells from anti-C1q positive donors were FACS sorted for anti-C1 q reactivity using C1q in solid phase. The cells were stained using C1q in complex with pre-formed IgG hexamers against DNP which were labelled with either PE or AlexaFluor647 and we sorted the double positive cells. Sorted cells were cultured and supernatant was screened for anti-C1 q reactivity, resulting in the successful identification of 9 unique anti-C1 q clones from a total of 4 healthy donors.

[0516] Variable domains of individual anti-C1q antibodies were sequenced and analyzed in IMGT V-quest to determine mutational load and V(D)J gene usage.

[0517] Anti-C1q mAbs were expressed recombinantly in the eukaryotic Expi293 expression system and purified for further analysis. Binding of the mAbs to C1q in ELISA varied greatly, with more than 100- fold difference in mAb concentration needed to saturate the plate between the strongest and weakest binding mAbs (Figure 1A). To exclude the possibility that the selected clones were in fact polyreactive, binding to several well-known targets of (polyreactive) autoantibodies was tested in ELISA. For all anti-C1 q mAbs, no meaningful binding to single-stranded DNA, lipopolysaccharide (LPS) or insulin was observed, demonstrating specificity of these mAbs to C1q (Figure 1 B). The binding affinity of isolated mAbs to C1q was also quantified by SPR, revealing single to double digit nanomolar avidity (Table 2).

[0518] Human monoclonal anti-C1q autoantibodies bind selectively to solid-phase C1q

[0519] Binding of mAbs to coated C1 q in the absence or presence of fluid-phase C1 q was analyzed by ELISA. Use of 20 pg / ml purified C1 q as a competitor did not yield any notable inhibition of mAb binding to solid-phase C1q, nor did the full C1qr2S2 complex (denoted C1 hereafter) as a competitor (Figures 2- 5). Adding C1 q-containing serum did not result in inhibition for the majority of mAbs. Only some of the mAbs, most notably mAb 4F5, were partially inhibited by C1q in high-concentration serum.

[0520] Finally, we studied the preferential binding of anti-C1q mAb to solid phase C1 q in great detail without any interference of bivalent binding or Fc-mediated C1 q interactions. For this purpose, we engineered antibodies with inactive Fc domains and containing only one C1 q-binding Fab arm for mAbs 1 F4, 1 F5 and 4F5. In ELISA, binding of these anti-C1q mAbs to coated C1q was analyzed in the presence of equal amounts of fluid-phase C1 q or C1q on beads (solid-phase). Binding was significantly inhibited only by C1q on beads, but importantly not by fluid-phase C1 q or human albumin, either on beads or in fluid-phase (Figure 6). Altogether, anti-C1 q mAbs show a strong selectivity towards binding solidphase, but not fluid-phase C1q.

[0521] Human monoclonal anti-C1q mAbs recognize C1q bound on a range of its natural ligands

[0522] Since solid-phase C1 q is also described as C1 q bound to its ligand, direct coating of C1 q to an ELISA plate may be seen as an artificial conformation. Therefore, binding of anti-C1q mAbs to C1q bound to a number of its natural ligands, such as IgG, IgM and CRP, was investigated in ELISA (Figure 7A). Binding of anti-C1 q mAbs to C1 q was detected on all of these ligands, providing evidence that antiCi q mAbs recognize C1q in its natural ligand-bound conformation. To examine the interaction of antiCi q mAbs with C1q in a natural solid-phase state in the context of a cell surface instead of an ELISA plate, binding on IgG opsonized cells and necrotic cells was measured (Figures 7B-C). Binding of the anti-C1 q mAb was specifically observed to alemtuzumab-opsonized cells in the presence, but not in the absence, of C1q. Antibody independent physiological C1q binding to dead cells was analyzed using necrotic PBMCs. C1q binding on necrotic cells was more modest as detected by the positive control polyclonal antibody. Nevertheless, several anti-C1 q mAbs showed binding to C1 q on necrotic cells, especially those identified in previous ELISA experiments as the strongest binding mAbs (Figure 1A, Figure 7C).

[0523] To investigate the importance of conformational structure of C1q for binding by anti-C1q mAbs, western blots loaded with C1q in different levels of unfolding (heated and reduced, heated only and unheated) were stained with anti-C1 q mAbs. Full blots shown as example demonstrate no background binding (Figure 8A). A strong staining of a band on the blot was observed on unheated C1 q for all antiCi q mAbs (Figure 8B). On heated samples of C1q, with or without reducing agent, none of the antiCi q mAbs yielded any signal. In contrast, a polyclonal antibody detecting C1 q showed clear and discrete bands at the expected molecular weights for all conditions. These results highlight the importance of the conformation of C1 q to allow anti-C1 q binding.

[0524] Human anti-C1q autoantibodies target multiple epitopes on the Collagen Like Region of C1q

[0525] To understand the nature and location of the epitopes that are targeted by these anti-C1 q mAbs, binding competition between mAbs was explored in ELISA (Figure 9A). While all anti-C1q mAbs display the expected self-inhibition, the mAbs show different levels of inhibition to the other mAbs. In fact, two distinct groups of mAbs competing with each other but not with mAbs from the other group could be discerned. Both groups consist of mAbs from multiple donors, while each group contains at least one mAb originating from donor 2, showcasing diversity even within one individual. This data shows that not all anti-C1q mAbs target the same epitope, and provides evidence for at least two different epitopes on C1q that are targeted by autoantibodies.

[0526] In order to map the binding of anti-C1q mAbs to the different regions of the C1q molecule, binding to CLR and recombinant globular head domain was evaluated (Figure 9B). All anti-C1q mAbs evidently bind to the CLR and not the globular head when tested in ELISA, with the exception of mAb 12F6, for which no targeted region could be identified.

[0527] The binding sites for the anti-C1q mAbs were compared with anti-C1q from SLE patients. To this end, antibodies from serum of three SLE patients (SLE A, B and C) were purified on protein A and competition for binding to C1q between mAbs and SLE antibodies was evaluated (Figure 14). Competition patterns were clearly split between the two groups identified in mAb-mAb competition. Epitopes targeted by SLE A and SLE B overlap with epitopes of mAbs 1 F4, 1 F5, 3C3 and 12F6, while SLE C antibodies compete with mAbs 2D2, 4C11 , 4D2, 4E6 and 4F5. Each anti-C1 q mAb was inhibited by the presence of at least one of the polyclonal SLE antibody mixes, showcasing that the anti-C1 q mAbs of the invention target the same or similar epitopes as anti-C1q autoantibodies in SLE patients.

[0528] In an effort to further specify the location on C1 q recognized by the anti-C1q mAbs, a set of 79 overlapping peptides was produced, with each peptide 21 amino acids long and 12 amino acids overlap between peptides, to cover all three chains of the C1q protein. Interestingly, no meaningful binding signal was observed for the anti-C1 q mAbs or anti-C1q positive plasma of 3 SLE patients on any of the peptides (Figure 12). As detection with a polyclonal anti-C1q antibody did show reactivity fora numberof peptides, the inability of the human anti-C1q autoantibodies to bind any of the peptides, together with the western blot data, points toward recognition of a structural epitope as opposed to an epitope consisting of a linear peptide.

[0529] Anti-C1q autoantibodies of the IgG isotype facilitate the recognition of opsonized particles by phagocytes

[0530] Although anti-C1q autoantibodies are often associated with disease, they are present in a portion of healthy individuals. Why healthy individuals harbor such autoantibodies is currently unknown, but we speculate that they may be triggered by and have a beneficial role in fighting infections. Possible advantages may include amplified opsonization of targets, leading to increased phagocytosis via Fc receptors. An experiment was set up to test binding of beads coated with IgM and C1 q to THP-1 cells differentiated into macrophages as a model for phagocytosis. In this setting, the binding or uptake of beads coated with only IgM and C1q is minimal, but addition of 1 pg / ml anti-C1 q mAb could increase binding of the beads to the differentiated THP-1 cells as much as 5-fold (Figure 10A). This enhancement of uptake could be inhibited by pre-incubating the cells with an Fc receptor blocking agent (Figure 10B). The combined data reveal that lgG1 anti-C1 q autoantibodies might improve the ability to clear particles opsonized by IgM by phagocytosis.

[0531] Electron tomography imaging ofanti-C1q

[0532] In order to obtain a molecular view on complexes of C1q with anti-C1q, we turned to electron tomography. Anti-C1 q binding was visualized on C1 / lgM complexes on liposomes with negative stain electron tomography, and on C1q / lgG complexes with cryo-electron tomography (Figure 11). As anti- C1 q antibody, bispecific Ab 1 F5xb12 (as also used in the solid-phase C1q inhibition ELISA) was employed, to be certain the anti-C1q mAb could not bind C1q through a second Fab arm or its Fc domain. After several clearly visible complexes were overlaid with a complex map to aid visual interpretation, two major findings could be identified. Firstly, the anti-C1q antibodies bind to C1q near the globular head region, and not the on the top of the C1 q molecule where all chains come together. Secondly, it is obvious that multiple anti-C1q antibodies bind to the same C1q protein, demonstrating the presence of multiple epitopes on one molecule. Although resolution was not high enough to pinpoint the exact location of interaction, these data provide an improved insight into the molecular interaction between C1q and anti-C1q antibody.

[0533] Discussion

[0534] In the above examples, the characteristics and functional properties of human anti-C1 q autoantibodies on a monoclonal level were investigated. These autoantibodies are present in several diseases such as SLE and HUVS but also in a substantial number of healthy individuals. Experimental studies indicate that anti-C1q autoantibodies contribute to renal disease only when there are C1q-containing immune complexes in the glomeruli (Trouw, J Clin Invest, 2004). In the above examples a total of 9 monoclonal antibodies were isolated from 4 normal donors, sequenced and recombinantly reproduced, these autoantibodies bind specifically to solid-phase C1 q, and not to C1q in fluid-phase. Solid-phase C1q in this study, could entail C1q bound to natural ligands like IgG, IgM and CRP, coated to ELISA plate wells or beads, or biotinylated C1 q on a streptavidin chip. Two separate groups of antiCi q mAbs recognizing different epitopes can be discriminated, displaying the limited heterogeneity of anti-C1 q autoantibodies.

[0535] Attempts by the inventors to isolate anti-C1q producing clones from SLE patients were not successful, likely because of the immunosuppressive treatment that the patients were taking.

[0536] As all anti-C1 q clones were first identified in ELISA with anti-IgG detection and amplified with primers specific for IgG, B cells producing these anti-C1q in vivo must have undergone class-switching. This notion is further reinforced by the high apparent affinity of anti-C1 q for its antigen, SPR revealed single to double digit nanomolar apparent affinities. However, the range of binding strength observed in ELISA and cellular assays was not mirrored in the SPR results, where little variation in apparent affinity was detected. Nevertheless, this difference may be explained by the fundamental difference in techniques, as SPR does not require interactions to resist multiple washing and incubation steps to be registered. These steps in ELISA and cellular assays may work to amplify any differences in binding strength.

[0537] In the above experiments, anti-C1 q binding to solid-phase C1 q was not or only marginally inhibited by fluid-phase C1q. Furthermore, anti-C1q binding could be inhibited in the same assay with solid-phase C1 q on beads, re-iterating the specificity of these antibodies. Anti-C1q binding was tested for C1q bound to natural ligands in ELISA, as well as IgG-opsonized cells and necrotic cells. Binding of antiCi q mAbs to C1 q on all of these ligands was detected. Binding to C1 q on necrotic cells could only be measured for some of the stronger binding anti-C1 q mAbs, probably due to a lower availability of C1 q on these cells.

[0538] On western blot, it was observed that anti-C1 q mAbs could only detect unheated C1q, the most structurally intact form tested. No binding was observed to heated or reduced C1 q, indicating binding to a structural rather than a linear epitope. The inventors have tried to map the epitope for the antiCi q mAb using linear overlapping peptides spanning the entire C1q molecule. However, despite a good signal for the polyclonal rabbit antibody response, no significant binding of the human sera or the mAbs was observed, which is in line with the notion that a conformational epitope is targeted.

[0539] To confirm that the analysis of anti-C1 q mAbs isolated from healthy donors is meaningful for the understanding of anti-C1q antibodies in SLE, the inventors performed competition experiments using purified IgG of anti-C1 q positive SLE patients. This confirmed that the anti-C1 q mAbs isolated from healthy donors indeed all target C1q epitopes that are also targeted by anti-C1 q autoantibodies present in SLE patients, underscoring the relevance of the identified anti-C1q mAbs.

[0540] In determining the epitope targeted by anti-C1q, the inventors could classify the anti-C1q mAbs in two groups based on competition assays. Furthermore, all but one mAb bound to C1 q CLR almost as strong as to full C1q. The binding to C1q CLR are in accordance with earlier studies on anti-C1q positive sera, which show anti-C1 q binding to the C1 q CLR (Wisnieski, J Immunol, 1992). The presented CryoEM images indicate binding of anti-C1 q mAb 1 F5 to an epitope on the CLR close to the globular head domain of C1q. While no exact epitope could be determined, this experiment contributes to identifying one of the regions of C1 q targeted by human anti-C1 q autoantibodies. The observation that multiple anti-C1q autoantibodies can bind to one C1q molecule also indicate that the target epitope is likely present on the extended arms of the C1q CLR and not on the central CLR where all arms come together. Collectively, the data indicate that anti-C1q is binding C1q in a conformationally changed state, a situation such as occurs in C1 q following binding to its natural ligands, or to surfaces.

[0541] Why healthy individuals would have high affinity anti-C1 q autoantibodies of the IgG subclass is completely unknown. However, once these antibodies have arisen, there may be some benefit under certain conditions. The inventors hypothesized that anti-C1 q of the IgG isotype could aid in phagocytosis of C1 q-opsonized particles by engaging Fc-receptors. Indeed, some anti-C1q mAbs increased binding or uptake of beads coated with IgM and C1 q up to 5-fold. In vivo, this could enhance the clearance of C1q-opsonized pathogens and apoptotic cells. In another situation, anti-C1q of complement-activating isotypes could enhance complement activation by depositing on target-bound C1 q. This is likely the mechanism by which deposited immune complexes in combination with antiCi q contribute to renal disease.

[0542] The above experiments are the first to describe monoclonal human anti-C1 q autoantibodies in great detail. The above experiments provide a detailed characterization of anti-C1q autoantibodies and allow to observe that multiple epitopes on C1 q can be targeted by anti-C1q. Focusing on the details from these monoclonal antibodies does not provide information on the relative abundance of each of the antibodies, but it is remarkable that antibodies with similar properties in all four donors were obtained. The competition analysis revealed two clusters of anti-C1q reactivity, consisting of mAbs from several donors. Collectively this indicates that the obtained clones may be representative of the anti-C1q IgG antibodies present in healthy individuals. By isolating only anti-C1q of the IgG isotype and producing it recombinantly in lgG1 subclass, some information on the presence of various isotypes and subclasses may have been lost.

[0543] The specific targeting of solid-phase C1 q could potentially be used diagnostically or therapeutically. In a diagnostic setting, the isolated anti-C1 q mAbs may be used to identify tissue locations where C1 q is activated in vivo. Importantly, anti-C1 q mAbs, or derivatives like bispecific antibodies, could also be deployed therapeutically to target locations where C1 q is activated. These antibodies would enhance complement activation through their Fc domain, for instance on cancer cells. In addition, antibodydrug conjugates may be developed to specifically target compounds to the locations where C1q is deposited.

[0544] Although the origin of anti-C1 q antibodies remains unclear, the current study provides insight into the molecular properties of human anti-C1q on a monoclonal level. These antibodies bind specifically to solid-phase C1q, which exposes cryptic epitopes not available in fluid-phase C1 q. These new recombinant antibodies may be developed for use in diagnostic tracing or modified for specific therapeutic targeting of solid phase C1 q.

[0545] Example 2

[0546] Materials and methods

[0547] ELISA for complement activation in presence ofanti-C1q mAb lgG1 anti-DNP with RGY mutations for hexamer formation were coated on ELISA plates at 10 pg / ml in coating buffer for 1 hour at 37 °C. The plates were blocked with PBS / 1%BSA for 1 hour at 37 °C, washed and incubated with 10 pg / ml C1q for 1 hour at 37 °C. After washing, 50 pg / ml anti-C1 q mAb was incubated in the wells for 1 hour at 37 °C. The wells were washed, incubated with 1 % NHS in RPMI 1640 (Gibco) for 1 hour at 37 °C and washed again. Deposited C5b-9 was detected using 333x diluted Mouse anti-C5b9 (clone aE11 , DAKO) and Goat anti-Mouse-HRP (DAKO). Plates were developed with ABTS / 0.015% H2O2 and absorbance at 415 nm was measured using a microplate reader.

[0548] ELISA for FcR 1 I la binding in presence of a nti-C1q mAb lgG1 anti-DNP with RGY mutations for hexamer formation were coated on ELISA plates at 10 pg / ml in coating buffer for 1 hour at 37 °C. The plates were blocked with PBS / 1%BSA for 1 hour at 37 °C, washed and incubated with 10 pg / ml C1q for 1 hour at 37 °C. After washing, 50 pg / ml anti-C1 q mAb was incubated in the wells for 1 hour at 37 °C. The wells were washed and then incubated with 3 pg / ml FcRIIIa-biotin for 1 hour at 37 °C. Detection of FcRIIIa-biotin after washing was performed by incubating with 0.1 pg / ml streptavidin-HRP for 1 hour at 37 °C. After adding ABTS / 0.015% H2O2, absorbance at 415 nm was measured using a microplate reader.

[0549] Binding ofanti-C1q opsonized beads by THP-1 cells

[0550] The THP-1 cell line (ATCC TIB-202) was cultured in RPMI (Gibco) with 10% FCS, 2mM L-glutamine, 100 units / ml penicillin and 100 pg / ml streptomycin (all Gibco). Cells were differentiated to a macrophage phenotype by incubating 100,000 cells / well in a 48 wells plate with phorbol 12-myristate 13-acetate (PMA; Sigma-Aldrich) in the medium. After 3 days incubating at 37 °C, medium was replaced with PMA-free medium and cells were incubated for another 5 days. Yellow fluorescent 0.46 pm carboxyl beads (Spherotech) were loaded with IgM for 1 hour at 37 °C, washed 3 times in PBS, and incubated with C1 q for 1 hour at 4 °C. After washing, beads suspension was incubated with antiCi q mAb in serum-free medium for 1 hour at 4 °C, after which the mixture was added to the differentiated THP-1 cells. Per well, 3 x 108beads, incubated with 0.1 pg IgM, 0.5 pg C1 q and 0.2 pg anti-C1 q mAb were added in a total volume of 200 pl serum-free medium. For experiments with Fc blocking reagent, cells were incubated with 10 pg / ml Human Fc block (BD Biosciences) for 10 minutes at room temperature before adding the beads suspension. Plates were centrifuged for 1 minute at 100 g to bring the beads in contact with the cells and were then incubated for 1 hour at 37 °C to allow binding and phagocytosis of anti-C1 q mAb-covered beads by the cells. The beads suspension was removed and the cells were treated with trypsin (Gibco) to detach them from the plate. Cells were resuspended, washed in FACS buffer and then analyzed on a FACSCanto flow cytometer. Binding or phagocytosis was measured by FITC fluorescence. Differentiation of THP-1 cells was confirmed by increased expression of CD11 b (by mouse anti-CD11 b-APC, clone D12, BD Biosciences) and decreased expression of CD15 (by mouse anti-CD15-BV510, clone W6D3, BD Biosciences).

[0551] Phagocytosis of S. aureus by human neutrophils

[0552] Human polymorphonuclear (PMN) leukocytes were isolated freshly from blood of healthy donors by the Ficoll-Histopaque gradient method. In 96-well plates, mAmetrine-labelled Staphylococcus aureus (strain NewmanAspa / sbi; 750000 cells / well) was mixed with 1 ,5 pg / ml monoclonal antibody (lgG4 with E430G mutation) against wall teichoic acid (WTA) and 6 pg / ml C1 q in RPMI supplemented with 0,05% HSA. Plates were incubated for 15 minutes at 37 °C on an orbital shaker. Subsequently, anti-C1q mAbs were added at 10 pg / ml and plates were incubated for another 15 minutes at 37 °C on an orbital shaker. Finally, PMN leukocytes cells were added at 75000 cells / well to allow phagocytosis of opsonized bacteria. After 15 minutes incubation at 37 °C, phagocytosis was stopped by addition of 1% paraformaldehyde. Phagocytosis was analyzed on a BD FACS Verse flow cytometer. Neutrophils were gated based on Forward / Sideward scatter and the fluorescence of mAmetrine-labelled bacteria associated with neutrophils was acquired.

[0553] Results

[0554] Anti-C1q mAbs increase Fc receptor engagement, but not complement activation, on immune complexes

[0555] The inventors sought to understand the consequences of the presence of anti-C1q autoantibodies on the activation of the immune system by C1 q-containing immune complexes. When anti-C1 q binds to C1 q on an immune complex, its Fc domain may add to the immune response by activating the complement system or by engaging Fc receptors (FcR). Plate-bound IgG complexes allow classical pathway complement activation, in this case detected as a dose dependent increase of C5b9 deposition with increasing serum concentration between 0.5% and 4% NHS (Figure 13A). From this titration the inventors selected 1 % NHS as a source of serum to analyze if the presence of anti-C1q mAb would impact on the degree of complement activation. The presence of anti-C1q mAb does not or only marginally increases complement activation (clones 1 F5 and 12F6), and in the case of 2 mAb (clones 3C3 and 4F5) a clear decrease of C5b9 was even observed (Figure 13B).

[0556] When investigating FcR engagement in the same context, deposited IgG complexes were bound by FcyRllla, and this binding was largely prevented when C1q was present on the hexameric IgG (Figure 13C). From the titration the inventors chose 3 pg / ml of FcRIII to analyze the impact of the anti-C1q mAb. The inventors observed that several anti-C1q mAbs were able to strongly increase FcyRllla engagement after binding to C1 q on immune complexes (Figure 13D). Focusing further on Fc receptor engagement by anti-C1 q mAbs, the inventors investigated binding or phagocytosis of IgM-coated beads opsonized with C1q by THP-1 cells differentiated towards macrophages. In this setting, the binding or uptake of lgM-coated / C1 q opsonized beads is minimal, but addition of 1 pg / ml anti-C1 q mAb increased binding of the beads to the differentiated THP-1 cells as much as 5-fold (Figure 13E). This enhancement of uptake was (partially) inhibited by pre-incubating the cells with an Fc receptor blocking agent (Figure 10B) confirming that this is an Fc receptor driven process. In a similar fashion, the inventors studied phagocytosis of S. aureus bacteria by PMN leukocytes when opsonized with anti-WTA lgG4. This lgG4 antibody contains mutation E430G for increased C1q binding, but does not result in increased phagocytosis by itself. Again, anti-C1 q mAb were able to enhance phagocytosis after opsonization with an antibody isotype that does not facilitate phagocytosis (Figure 13F). Anti-C 1 q autoantibodies may thus increase Fc receptor engagement in vivo, in places where immune complexes with C1q have deposited. Thus, in vivo, anti-C1 q antibodies, such as those according to the present invention, could enhance the elimination of C1 q-opsonized objects such as pathogens, apoptotic cells, and cancer cells via FcR expressing (immune) cells. This may not be necessarily limited to phagocytosis, but could also comprise other FcR-mediated effects, for example antibodydependent cell-mediated cytotoxicity. Alternatively, anti-C1q antibodies, such as those according to the present invention, comprising modifications that inhibit their binding to FcR, for example in the Fc domain, could ameliorate FcR-mediated immune cell activity, which may be beneficial during for example the treatment of auto-immune disease or organ transplantation.

Claims

Claims1. An isolated polypeptide comprising an antibody variable domain that specifically binds to human solid-phase complement component C1q.

2. The polypeptide according to claim 1 , wherein the polypeptide is an isolated antibody or antigen-binding fragment thereof.

3. The polypeptide according to claim 1 or 2, wherein the polypeptide comprises an antibody variable domain that specifically binds to human solid-phase complement component C1q in that in an in vitro competitive binding assay the polypeptide exhibits less than 25% inhibition in binding to solid-phase C1 q in the presence of an excess of fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1 q in the absence of fluid-phase C1 q, preferably wherein the excess of fluid-phase C1q is 20 pg / ml purified fluid-phase C1q, the solid-phase C1 q is provided at a concentration of 10 pg / ml, and wherein the degree of inhibition is determined at or near the saturation concentration of the polypeptide when binding to solidphase C1q in the absence of fluid-phase C1q.

4. The polypeptide according to claim 3, wherein the in vitro competitive binding assay comprises an ELISA Competition Assay and wherein in the ELISA Competition Assay, the polypeptide exhibits less than 25% inhibition in binding of solid-phase C1q in the presence of 20 pg / ml purified fluid-phase C1q, compared to binding of the polypeptide to solid-phase C1q in the absence of fluid-phase C1q, preferably wherein the solid-phase C1q is provided by coating an ELISA plate surface with 10 pg / ml C1q, and wherein the degree of inhibition is determined at or near the saturation concentration of the polypeptide when binding to solid-phase C1q in the absence of fluid-phase C1q.

5. The polypeptide according to any one of claims 1 to 4, wherein the polypeptide has a dissociation constant (KD) for solid-phase C1 q of less than about 3.0 x 10'8M, preferably of less than about 2.5 x 10'8M, wherein the KD is determined by surface plasmon resonance.

6. The polypeptide according to claim any one of claims 1 to 5, wherein the antibody variable domain comprises: a. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 1 ;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 2;(iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 3; anda light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 4;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 5;(iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 6; or b. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 7;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 8;(iii) a CDR-H3 comprising an amino acid sequence having at least 92% sequence identity to SEQ ID NO: 9; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 10;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 11 ;(iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 12; or c. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 13;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 14;(iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 15; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 16;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 5;(iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 17; or d. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 18;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 19;(iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 20; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 22;(iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 23; or e. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 24;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 25;(iii) a CDR-H3 comprising an amino acid sequence having at least 96% sequence identity to SEQ ID NO: 26; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 21 ;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 22;(iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 27; or f. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 28;(ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 29;(iii) a CDR-H3 comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 30; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 31 ;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 32;(iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 33; or g. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 34;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 35;(iii) a CDR-H3 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 36; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 38;(iii) a CDR-L3 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 39; or h. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 40;(ii) a CDR-H2 comprising an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 41 ;(iii) a CDR-H3 comprising an amino acid sequence having at least 93% sequence identity to SEQ ID NO: 42; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 88% sequence identity to SEQ ID NO: 43;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 44;(iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45; or i. a heavy chain variable domain (VH) comprising(i) a CDR-H1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 46;(ii) a CDR-H2 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 47;(iii) a CDR-H3 comprising an amino acid sequence having at least 94% sequence identity to SEQ ID NO: 48; and a light chain variable domain (VL) comprising(i) a CDR-L1 comprising an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 49;(ii) a CDR-L2 comprising an amino acid sequence having at least 66% sequence identity to SEQ ID NO: 50;(iii) a CDR-L3 comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 51.

7. The polypeptide according to claim 6, wherein: a. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 1 , 2, and 3; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 4, 5, and 6; or b. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 7, 8, and 9; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 10, 11 , and 12; or c. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 13, 14, and 15; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 16, 5, and 17; or d. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 18, 19, and 20; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 21 , 22, and 23; or e. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 24, 25, and 26; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 21 , 22, and 27; or f. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 28, 29, and 30; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 31 , 32, and 33; or g. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 34, 35, and 36; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 37, 38, and 39; or h. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 40, 41 , and 42; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 43, 44, and 45; or i. CDR-H1 , CDR-H2, and CDR-H3 comprise the amino acid sequences of respectively SEQ ID NOs: 46, 47, and 48; and wherein CDR-L1 , CDR-L2, and CDR-L3 comprise the amino acid sequences of respectively SEQ ID NOs: 49, 50, and 51 .

8. A conjugate comprising a polypeptide according to any of claims 1 to 7 conjugated to or recombinantly fused to a diagnostic agent, detectable agent, or therapeutic agent.

9. The polypeptide or conjugate according to any of claims 1 to 8, wherein the polypeptide is a human antibody.

10. The polypeptide or conjugate according to any of claims 1 to 9, wherein the polypeptide is selected from the group consisting of a: whole antibody, bispecific antibody, diabody, triabody, tetrabody, minibody, nanobody, sdAb, scFv, di-scFv, bis-scFv, tri-scFv, scFv-Fc, Fab, Fab', F(ab')2, and Fv.

11. The polypeptide or conjugate according to any of claims 1 to 10, wherein the polypeptide is a monoclonal antibody.

12. The polypeptide or conjugate according to any of claims 6 to 11 , wherein the polypeptide further comprises a heavy chain constant region, optionally wherein the heavy chain constant region is selected from the group consisting of: IgG, IgM, IgE, IgD and IgA, further optionally wherein the heavy chain constant region is selected from the group consisting of: lgA1 , lgA2, lgG1 , lgG2, lgG3 and lgG4.

13. The polypeptide or conjugate according to any of claims 9 to 12, wherein the polypeptide comprises a modified Fc domain.

14. The polypeptide or conjugate according to claim 13, wherein the modified Fc domain has increased or reduced interaction with an Fc receptor compared to an unmodified Fc domain of the polypeptide.

15. The polypeptide or conjugate according to claim 13 or 14, wherein the modified Fc domain comprises an amino acid mutation, substitution, insertion, deletion, or addition, a modification to a glycan structure, a glycosylation site, or number of glycosylation sites, or a post- translational chemical, biochemical, or enzymatic modification.

16. An isolated nucleic acid comprising a sequence encoding a polypeptide or a conjugate according to any of claims 1 to 15, preferably wherein the sequence comprises a nucleic acid sequence encoding a modified Fc domain, preferably wherein the nucleic acid sequence encoding the modified Fc domain comprises a mutation, substitution, insertion, deletion, or addition of one or more nucleic acids of the sequence.

17. An isolated cell comprising a nucleic acid according to claim 16.

18. A pharmaceutical composition comprising a polypeptide or conjugate according to any of claims 1 to 15, and a pharmaceutically acceptable excipient, adjuvant, diluent, or carrier.

19. A kit comprising a polypeptide or conjugate according to any of claims 1 to 15, and instructions for using the polypeptide or conjugate to (i) treat or prevent infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, autoimmune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in a subject in need of such treatment, or (ii) diagnose or predict the development of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in vivo in a subject.

20. A polypeptide or conjugate according to any of claims 1 to 15, or a pharmaceutical composition according to claim 18, for use as a medicament or a diagnostic.21 . A polypeptide or conjugate according to any of claims 1 to 15, ora pharmaceutical composition according to claim 18, for use in the treatment of a disease or disorder associated with complement activation involving C1q.

22. A polypeptide or conjugate according to any of claims 1 to 15, ora pharmaceutical composition according to claim 18, for use in the treatment of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, auto-immune disease or cancer.

23. A polypeptide or conjugate according to any of claims 1 to 15, or a pharmaceutical composition according to claim 18, for use in immunotherapy or transplantation, preferably wherein the use in transplantation comprises use in the pre-treatment of a subject prior to receiving transplantation of an organ, tissue or cells, use in the pre-treatment of an organ, tissue or cells prior to transplantation to a recipient subject, or use in the treatment or prevention of tissue or organ rejection in a subject.

24. A polypeptide or conjugate according to any of claims 1 to 15, or a pharmaceutical composition according to claim 18, for use in a method of diagnosis or prediction of development of infectious disease, neurological disease, neurodegenerative disease, graft versus host disease, tissue or organ rejection, auto-immune disease, cancer, or a disease or disorder associated with complement activation involving C1q, in vivo in a subject.

25. A polypeptide, conjugate, or pharmaceutical composition for use according to claim 20 to 24, wherein the polypeptide, conjugate, or pharmaceutical composition is formulated for use with one or more other pharmaceutical compositions or one or more diagnostic agents, detectable agents or therapeutic agents, preferably wherein the one or more other pharmaceutical compositions or one or more diagnostic agents, detectable agents or therapeutic agents is selected from the group consisting of a: complement activating compound, complement inhibiting compound, tracer, label, radioactive compound, toxin, CRISPR-associated protein(Cas), preferably wherein the Cas is Cas9, and nucleic acid, preferably wherein the nucleic acid is a ribonucleic acid (RNA) or a deoxyribonucleic acid (DNA), more preferably wherein the RNA is selected from the group consisting of a: small interfering RNA (siRNA), asymmetrical interfering RNA (aiRNA), microRNA (miRNA), Dicer-substrate RNA (dsRNA), small hairpin RNA (shRNA), messenger RNA (mRNA), antisense RNA (asRNA), aptamer, circular RNA (circRNA), self-amplifying RNA (saRNA), catalytic RNA, anti-miRNA, long noncoding RNA (IncRNA), single-guide RNA (sgRNA) and mRNA encoding a Cas, or wherein the DNA is selected from the group consisting of a: antisense oligonucleotide (ASO), aptamer, episome and catalytic DNA.

26. Use of a polypeptide or conjugate according to any of claims 1 to 15 for detecting solid-phase C1 q.

27. A method of detecting solid-phase C1q in a biological sample, comprising the steps of: a. contacting a polypeptide or conjugate according to any of claims 1 to 15 with the biological sample; and b. detecting the presence or absence of the polypeptide bound to solid-phase C1 q in the biological sample.

28. A method of detecting solid-phase C1q in vivo in a subject, comprising the steps of: a. administering a polypeptide or conjugate according to any of claims 1 to 15, or a pharmaceutical composition according to claim 18, to the subject; and b. detecting the presence or absence of the polypeptide bound to solid-phase C1 q in the subject.

29. A method of removing C1q in vivo in a subject, comprising the steps of: a. administering a polypeptide or conjugate according to any of claims 1 to 15 or a pharmaceutical composition according to claim 18, to the subject; and b. removing solid-phase C1 q or complexes comprising solid-phase C1 q from a target tissue to which they are bound.

30. A method of treating a subject, comprising administering a therapeutically effective amount of a pharmaceutical composition according to claim 18 to a subject in need thereof.31 . A polypeptide or conjugate according to any of claims 1 to 15, or a pharmaceutical composition according to claim 18, for use in the treatment of systemic lupus erythematosus (SLE).

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